Validity of echocardiographic estimates of left ventricular size and performance in infants and children. Echocardiography has been shown to be a reliable method for estimating left ventricular size and function in adults, but little attention has been paid to its application to infants and children. This paper describes a validatory study in 40 children aged between 4 days and 16 years. There was a significant correlation between angiocardiographic and echocardiographic estimates of left ventricular end-diastolic volume (r=0.76), left ventricular end-systolic volume (r=0.68), and ejection fraction (r=0.73). Left ventricular mean circumferential shortening rate (mean Vcf), however, gave the mostsignificant correlation (r=0.91). These findings indicate that while care must be exercised in interpreting indices of left ventricular size and performance derived from echocardiography, these measurements, and in particular mean Vcf, do offer a useful and reliable means of assessing left ventricular function in infants and children.


Accuracy of echocardiography for assessing aortic root diameter. The normal range for aortic root diameters employing the echocardiographic continuous recording technique was determined in 159 adult subjects without aortic valve disease or hypertension. In order to evaluate the accuracy of this noninvasive technique, the aortic root diameters as measured by ultrasound before operation in 31 patients with aortic valve disease were compared with their respective aortic annulus diameters as determined at the time of valve replacement. The 159 normal subjects consisted of 78 men and 81 women whose ages ranged from 18 to 74 pears (average 38). The normal range of aortic root diameters in this group was 17 to 33 mm (mean 23.7). A significant difference (P is smaller than 0.001) in aortic root diameters existed between men and women which could not be explained by differences in body surface area. The mean diameter among the 81 women was 22.4 plus or minus 3.3 mm(I SD) compared to the mean diameter in the 78 men of 25 plus or minus 3.5 mm. These results indicate a normal range for aortic root diameters which is significantly lower than previously reported. The aortic root diameter which was measured reoperatively by ultrasound in the 31 patients with aortic valve disease averaged 25.1 plus or minus 1.9 mm and did not differ significantly from the actual aortic annulus diameter of 25.5 plus or minus 2.6 mm measured at the time of surgery. The aortic root measurements by echo were within 2 mm of the corresponding annulus diameter in 25 of the 31 operated patients (80%).


Comparison of scalar and vector electrocardiographic diagnosis and localization of myocardial infarction. A prospective study was made of 80 patients during typical clinical episodes of acute myocardial infarction with biochemical and scalar electrocardiographic confirmation. Nine patients had bundle-branch block and 12 had had previous episodes of myocardial infarction. Serial electrocardiograms and vectorcardiograms were recorded during the first week in hospital. The most striking finding was that in more than half the cases there was disagreement between the electrocardiogram and vectorcardiogram in the localizationof infarction. The earliest evidence of infarction in the electrocardiogram is often restricted to ST and T changes though in the corresponding vectorcardiograms evidence of infarction may be present in the QRS loop. In those cases in which both electrocardiogram and vectorcardiogram show QRS abnormalities these are frequently detected earlier in the vectorcardiogram, but the converse is rare.


Right-to-left interatrial shunting with normal right ventricular pressure. A puzzling haemodynamic picture associated with some rare congenital malformations of the right ventricle and tricuspid valve. Haemodynamic and angiographic findings are described in 5 cases with rare anomalies of the right ventricle and the second there was isolated congenital 'absence' of the tricuspid valve. The remaining 3 cases all had isolated congenital hypoplasia of the right ventricle; in 2 of these 3 cases closure of the atrial septal defect was successful in abolishing cyanosis and symptoms,while the third is awaiting surgical correction.


Left and right heart haemodynamics during spontaneous angina pectoris. Comparison between angina with ST segment depression and angina with ST segment elevation. The function of both right and left sides of the heart was studied during spontaneous attacks of angina pectoris at rest in 7 patients showing ST depression (type I) and 4 showing ST elevation (type II) during the attack. In none of the 44 type I attacks and 29 type II attacks which were recorded did circulatory changes; the latter were different in the two groups. Type I attacks showed: a) a brief fall in arterial pressure, accompanied by b) a rise of right atrial and pulmonary wedge pressures and c) a decrease of cardiac output, right and left stroke work, the mean rate of systolic ejection, and indirect left ventricular pre-ejection dP/dt. In the course of the attack a hypertensive phase followed, which was paralleled by an increase of heart rate, cardiac output, left and right stroke work, and mean systolic ejection rate, left dP/dt; right atrial pressure and wedge pressure remained raised. All of the circulatory functions started to revert towards the pre-attack levels coincident with the waning phase of the electrocardiographic alteration, the latter occurring either spontaneously or after nitroglycerin. Type II attacks for the entire duration of the electrocardiographic changes showed: a) a reduction of arterial pressure, cardiac output, right and left stroke work, mean systolic ejection rate, and left dP/dt, b) a rise of right atrial and wedge pressures, and c) quite small changes of heart rate. When the electrocardiogram started to revert to the pre-attack aspect, the cardiac function rapidly improved and, after a supernormal phase, returned to the basal levels in about 2 minutes. It is concluded: 1) that no circulatory factor interfering with the mechanical effort of the heart is responsible for eliciting spontaneous angina: 2) that in type I attacks right and left ventricular impairment occurs which recovers rapidly, possibly through a sympathetic compensation; 3) that in type II attachs dysfunction of both sides of the heart occurs and persists throughout the episode of electrocardiographic alteration; 4) that the dynamic impairment is probably more severe in type I than in type II angina.


Sinus node recovery time in the elderly. Measurement of the sinus node recovery time has been proposed as a diagnostic tool for recognition of the sick sinus syndrome. The latter is most frequently encountered in elderly patients with hypertension, coronary heart disease, and atherosclerosis. In order to provide normal values for the sinus node recovery time in this particular population group, atrial pacing studies were carried out in 30 subjects over 50 years of age, all with peripheral vascular disease and some with angina pectoris (10), residua of infarction (6), or hypertension (7). On stimulation, 7 patients maintained a I:I atrioventricular conduction up to the rate of 180/min. Second degree atrioventricular block developed in all other cases. On six occasions, Wenckebach's periods appeared at the relatively slow pacing rate of 120/min. The maximum postoverdrive pause ranged from 680 to 1600 ms with an average of 1100 ms plus or minus 190 (10). For each pacing speed, a correlation was found between the duration of the pause and the control intrinsic cardiac rate, longer pauses being associated with longer resting PP intervals. Beyond 120/min, the duration of the pause was seen to shorten progressively as the driving rate was increased. Finally, the behavior of the sinus node pacemaker following interruption of pacing showed individual variations. After pacing at relatively slow rates, a prompt return to near control values was consistently observed, whereas, after fast rates of driving, a phase of secondary depression developed in about one-half of the studied cases.


Effects of the quaternary ammonium compound OX-572 on ventricular tachyarrhythmias complicating acute myocardial infarction. Patients with acute myocardial infarction who developed defined ventricular tachyarrhythmias were randomly allocated to two groups. The patients (n=34) in one of the groups received the quaternary ammonium compound N,N-bis (phenylcarbamoylmethyl) dimethylammoniumchloride (QX-572) intravenously as an infusion of 8 mg/kg body weight over 30 minutes. The patients in the other group (n=33) received saline and acted as controls. The analysis of arrhythmias was based on a continuous electrocardiogram recorded in parallel to routine monitoring during the 24 hours after the start of infusion. Only the detection of ventricular tachycardia or the development of ventricular fibrillation on routine monitoring could induce a change in antiarrhythmic treatment. The incidence of patients with ventricular tachycardia recorded was significantly lower in the QX-572 group (38%) than in the control group (73%). About half the number of patients with ventricular tachycardia recorded on the continuous electrocardiogram were also detected by means of routine monitoring. The number of patients with various types of ventricular premature contractions (1-5/min, greater than 5/min, paired, multifocal, R on T), ventricular tachycardia, and rapid idioventricular rhythm were compared within successive one-hour periods. For all types of ventricular tachyarrhythmias except rapid idioventricular rhythm there was a lower incidence of arrhythmias in the QX-572 group...


Anomalous origin of left coronary artery from pulmonary artery. Surgical considerations in the adult. The usually recommended treatment for anomalous origin of a left coronary artery from the pulmonary artery is proximal ligation of the anomalous artery with a saphenous vein bypass graft to restore aortocoronary continuity. In an adult patient with large collateral vessels and with the left coronary artery arising from the back of the pulmonary artery, the technical surgical problems associated with this procedure are formidable. Closure of the orifice of the anomalous left coronary artery from inside the pulmonary artery is suggested as the treatment of choice in such a case.


Changing haemodynamics in patient with papillary muscle dysfunction. A patient with papillary muscle disease caused by myocardial infarction was studied before and after injection of phenylephrine. The pulmonary wedge pressure was normal at rest. However, pressures and murmur changes, occurring spontaneously and after injection of phenylephrine, suggested that intermittent severe mitral regurgitation contributed significantly to the recurrent episodes of acute left heart failure presented by this patient.


A follow-up study of coronary heart disease in viscose rayon workers exposed to carbon disulphide. In 1967 two cohorts of 343 men each were formed and matched with respect to age, district of birth, and similarity of work. One cohort comprised viscose rayon workers with at least five years' exposure to carbon disulphide during any period between 1942 and 1967, and the other cohort consisted of workers from a paper-mill with no such exposure. The concentrations of carbon disulphide and hydrogen sulphide in the workroom air had been measured regularly since 1950, and about 4000 measurements were available. In all probability the concentrations had been very high in the 1940s, between 20 and 40 ppm in the 1950s, and about 10 to 30 ppm from 1960 onwards. On examination in 1967 it was confirmed that all of the relevant coronary risk factors had been kept under control. The only exception to this was blood pressure which was slightly higher among the exposed workers, a finding that was interpreted as a result of exposure rather than an independent risk factor. A five-year follow-up showed that 14 men had died from coronary heart disease (CHD) in the exposed group, against three in the control group (P smaller than 0:007). Other causes of death were evenly distributed. In addition, 11 nonfatal first infarctions had occurred in the exposed group as compared with four in the control group. On re-examination in 1972, nearly 25% of the exposed men, against 13% of the controls, had a history of angina (typical, probable, and possible) as measured by the  World Health Organization questionnaire (P smaller than 0:0002). The prevalence proportions of typical angina were 12% and 5% respectively (P smaller than 0:001). As opposed to this, only slight differences were apparent for coronary ECGs. The higher degrees of prevalance in the exposed group may well have been attributable to chance. As in 1967/68, the mean systolic and diastolic blood pressures were slightly higher in the exposed group (P smaller than 0:001 and P smaller than 0:01, respectively). The relative risk was 4-8 for fatal attacks, 3-7 for all infarctions, 2-8 for nonfatal infarctions, 2-2 for angina, and 1-4 for ECG findings indicative of CHD. This implies that with increasing severity and specificity of the manifestations the causal role of CS2 in developing CHD becomes more evident. Further, it is inferred that exposure to CS2 seems to worsen the prognosis of CHD in addition to increasing its incidence. Although the exposure data in this study may not be representative of the personal exposure of the workers, the conclusion is drawn that a great deal of justification exists for lowering the threshold limit value of 20 ppm recommended by the American Conference of Governmental Industrial Hygienists.


Thenar muscle blood flow and bone mineral in the forearms of lumberjacks. Forty lumberjacks who had used a chain saw for 0-20 years and who had no general disease affecting the bones were studied by measuring the thenar muscle blood flow of both hands by the 133-Xe local clearance method. Bone mineral in the left forearm in the region of cancellous and cortical bone was assessed by the 241-Am gamma ray attenuation method. Virbration was found to decrease the blood flow in the saw-bearing left hand compared with the right hand of the lumberjacks. The bone mineral density (g/vm-3) was lower in the forearm bones of the lumberjacks than in controls of the same age with healthy bones. Moreover the poorer the thenar muscle blood flow, the greater was the decrease in the mineral density of the distal radius. Measurement of the mineral density of the forearm bones by the gramma ray attenuation method can be used for early detection of bone lesions in traumatic vasospastic disease.


Prevalence of asbestos bodies in a necropsy series in East London: association with disease, occupation, and domiciliary address. The prevalence of asbestos bodies was measured in lung sections in a necropsy series carried out at the London Hospital (1965-66) after exclusion of all known asbestos factory workers and cases of asbestosis and of mesothelioma. Associations were sought between the presence and number of asbestos bodies with the patients' sex, domiciliary address, occupation, industry, and diseases recorded at necropsy. Asbestos bodies were present in 42% of the 216 men in the series and in 30% of the 178 women. The number of bodies in the positive cases was small in comparison with the numbers seen typically in asbestosis; thus there were less than 6 asbestos bodies per 6-75 mm-3 lung tissue in 107 of the total 145 positive cases in contrast to 1 000 or more in asbestosis. In comparison with the overall series, an increased number of asbestos body positives was present in males with carcinoma of stomach and females with carcinoma of breast. In view of this finding lung sections were counted in further post-mortem examples of these carcinomas making a total of 50 males with carcinoma stomach and 82 females with carcinoma breast. Thirty-five positive cases were found in the carcinoma stomach group as against 22-7 expected and 38 in the carcinoma breast group against 26-35 expected. There was no excess of observed over expected asbestos body positives in 51 males with carcinoma of bronchus. There was an excess of asbestos body positives (60-9%) in heavy manual workers and in both heavy and light manual male workers in the shipping (61%), electrical and engineering  (56%), and transport (54%) industries. The incidence in male clerical workers was 12-8%. The incidence of asbestos body positives according to home address was highest (53% in males, 45% in females) in patients living in the industrial and cockland area due east of the hospital. The incidence fell in the less industrial areas north-east of the hospital. Consideration of possible environmental sources of the inhaled asbestos suggests that in this survey occupation, industry, and comiciliary area all play a part. The comparatively minor intensity of asbestos pollution in our positive cases showed a positive association with carcinoma of stomach and breast, possibly playing a direct pathogenic role in carcinoma of stomach. No positive association was identified with any other neoplastic disease including carcinoma of bronchus.


Effect of asbestos on lipid peroxidation in the red cells. In vitro exposure of red cells to vie International Union against Cancer (UICC) standard reference asbestos samples resulted in an increase of thiobarbituric acid substances. Chrysotiles developed the largest amounts of lipid peroxides, followed by anthophyllite, amosite, and crocidolite in decreasing order. Compared with the control samples erythrocytes free of dusts, all types of the asbestos examined disclosed significant differences. The results obtained provide support for the cytotoxic potential of amosite and crocidolite and, on the other hand, suggest that a lipid peroxidation of unsaturated fatty acids may be involved in the mechanisms(s) of membrane-damaging effects of asbestos dusts.


Effect of repeated exposure to aniline, nitrobenzene, and benzene on liver microsomal metabolism in the rat. Exposure of rats to aniline at daily doses of 50 mg/kg of body weight over a month stimulated the microsomal metabolism as manifested by (1) acceleration of p-hydroxylation of anilin and N-demethylation of aminopyrine in 9-000 times g postmitochondrial supernatant of the liver, (2) shortening the sleeping time after hexobarbital, and (3) reduction of the antipyretic effect of phenacetin. In the rats exposed to nitrobenzene in a similar manner to aniline, nitroreduction of nitrobenzene and p-hydroxylation of aniline remained unaffected; the antipyretic  effect of phenacetin was decreased, whereas hexobarbital sleeping time remained unchanged. Exposure of rats to benzene (50 mg/kg of body weight daily for a month) had no effect on the rate of hydroxylation of benzene and N-demethylation of aminopyrine. In benzene-exposed rats hexobarbital sleeping time was prolonged whereas the antipyretic effect of phenacetin was  unaffected. Microsomal metabolism of aniline, nitrobenzene, and benzene was stimulated and inhibited when the rats were pretreated with phenobarbital and SKF 525-A, respectively.


Haemolytic episode in G6 PD deficient workers exposed to TNT. This is a report on three cases of acute haemolytic disease in glucose-6-phosphate dehydrogenase (G6PD) deficient workers exposed to trinitroluene (TNT). The courses of the haemolytic crises have several features in common, the most striking being the onset of the disease within two to four days after the start of exposure, and the fact that this has been the first and so far the only haemolytic episode in their lives in spite of detailed medical records of past diseases, injuries, and medications during 12 to 16 years preceding the haemolytic crisis and five to nine years following the disease. The lowest haemoglobin levels for the three patients were 4-0, 6-8, and 8-2 g/dl respectively; haematocrit values were 17 and 24%; reticulocytes rose in case 1 to 26-2%, in case 2 to 26%, and in case 3 to 10%. Indirect bilirubinaemia was increased in two patients (5-1 and 2-6 mg/100 ml) and stercobilinogen was as high as 2150 mg/24 hr in one patient. The presence of the metabolite monoamino 2-6 dinitrotoluene was proved in the urine of case 3 and reached 21 gamma%. The possibility of a dose-response relationship is briefly discussed and the risk of exposing G6 PD individuals in chemical processes is mentioned.


A clinical and radiographic study of coir workers. Processing of coir, which is the fibre obtained from the husk of the coconut, is a dusty procedure; 779 workers in two coir processing factories in Sri Lanka were examined clincally and radiographically for evidence of respiratory disease. Respiratory symptoms were present in 20 (2-6%) of them, which is no higher than in the general population. Respiratory disease such as asthma, chronic bronchitis, byssinosis, and pulmonary tuberculosis which may occur from occupational exposures were considered, but there was no evidence to suggest a definite association between these conditions and coir dust. Twenty-two workers had abnormal chest radiographs, but when compared with a control group of 591 workers from an engineering firm where lesions were found in 20 cases, there was no significant difference. In the opinion of the medical officer, management and workers of the large factory investigated, coir dust does not produce any respiratory disability. The chemical composition of coir dust is similar to that of sisal which is also relatively inert.


The medical school and smoking.

The medical school and smoking. To examine the social environment of the medical school as it relates to smoking, tape-recorded interviews were conducted with 92 medical students at the University of Manchester. The medical school provides a social setting unfavourable to smoking for many students, but for some non-smokers and many smokers countervailing forces lead to an expansion of the habit during training. Increased opportunity, the influence of peers and social factors, together with environmental stress, tend to encourage smoking. The necessary information about smoking is provided and this influences some, but in general the medical school fails to establish a social climate in which smokers would feel more strongly motivated to give up. Although most students accept in theory that non smoking is part of the doctor's role, this idea is not consistently presented to them during the course. Furthermore, it is not always seen in practice to be adhered to by members of staff, their role models, thus enabling the student who smokes to deny the relevance of his personal behaviour to his future role. Nor is this ameliorated by the students' perception of policy in the medical school as one of, at most, mild disapproval.


Effect of "attitudinal content" on formation of professional attitudes in medical students. The findings of a previous paper by the authors published in the British Journal of Medical Education (1974, vol. 8, p. 13) are summarized. Its significance as a foundation for this report is developed. The nature of the attitudes under study, the methods and the analytical techniques employed in the analysis are discussed briefly. The proposition to be examined involves the effect of the attitudinal content as a predetermining factor in the formation of professional attitudes in medical students. The distribution of attitude scale scores for the three attitudes examined are reported tabularly for the faculty by career activity and specialty, and for students in their aspired-to career activity and specialty. The analysis conducted evaluates the relative importance of socialization, generational and self-selection factors in the development of professional attitudes in medical students, The findings indicate that generational effects are present with all three attitudes, that self-selection is operative in two, and socialization appears only to occur with regard to one of the attitudes.


Use of a programmed reinforcement exercise in teaching of embryology. In studying embryology, understanding of the initial complexities of cell division and patterns of prenatal growth demands ability in three-dimensional comprehension. A programmed reinforcement exercise (PRE) was designed to enable second year medical students to devote their complete attention to the explanatory component of lectures, to highlight salient features in lectures, and to improve the efficiency and active participation component of revision. The programmed reinforcement exercise was used in association with a concurrent programme in gross anatomy and its effectiveness was evaluated by a controlled trial in two consecutive lectures to 142 students. With the t-test and matched pair tabular analysis to assess results, students using the programmed reinforcement exercise achieved significantly higher scores in two 15-item multiple choice tests which were designed to appraise their short-term comprehension and data recall of lecture content.


Effects of a short educational planning workshop on attitudes of three groups of medical educators. A series of short (3 days) educational planning workshops was held in three African faculties of medicine. The purpose of this study was to ascertain the effect of such a workshop on educational attitudes of teachers from three conservative traditional schools. An attitudes questionnaire given before and after the workshop allowed the participants to express opinions and feelings on a number of statements with a bipolar theme of teacher-orientation vs. student-orientation. The results showed that using experiential learning methods in small groups with little or no didactic presentation, it was possible in a short time to change long-held educational views of diverse groups of medical educator.


Medical sociology in Great Britain.

Medical sociology in Great Britain. The paper reports on a survey into the present state of medical sociology in Great Britain, describing the developments which have occurred following upon the recommendations in the Todd Report on the teaching of behavioural sciences to undergraduates. Details of existing and proposed courses at each school are given; they reveal the wide variations which exist, in respect of precise specification of educational objectives, course content, affiliation and qualifications of teachers, teaching methods, and curricular time. However, most schools are consciously trying to incorporate this discipline into the syllabus at an early stage. At present several of the provincial schools have more experience of teaching medical sociology than do the London schools, but this difference may soon be lessened. In most cases the subject is taught in close association with social or community medicine, but connexions with psychology and psychiatry are often close. The prospects for this relatively untried subject are good, with new medical schools demonstrating new ways of applying sociology to medicine.


Development and evaluation of teaching course in general practice. The further development of a teaching course in general practice medicine for fifth year students at Glasgow University is described. Lessons learnt from a previous experimental course have been applied: eight afternoon sessions of a structured nature and related to prescribed objectives and a syllabus were given to small groups of students. Questionnaires showed that the in-depth study of selected patients in their own homes, with student involvement in the preparation of problem lists for group discussion were thought by the students to be valuable. An objective assessment of the course using the modified essay question technique showed an improvement in factual clinical knowledge (P less than 0.05) and more importantly in areas relating to the general practice content of the teaching (P less than 0.001).


Postgraduate degree course in clinical biochemistry. A report on the progress of the M.Sc. courses in clinical biochemistry run at the University of Surrey over the past 7 years is given. The previous experience and qualifications of the students and their careers subsequent to taking the course are analysed. A description of the course structure and syllabus is given. It is concluded that the course has been useful in attracting both science graduates and medical graduates of high calibre, in providing them with a sound laboratory-based training in clinical biochemistry, and in stimulating their interest in following a career in clinical biochemistry and chemical pathology.


Young psychiatrists.

Young psychiatrists. Psychiatry is failing to recruit sufficient British graduates, reflecting a decline over the past 10 years in the proportion of medical students who are contemplating a career in psychiatry, and there are indications that the specialty is not getting its fair share of the best graduates. By a further analysis of the 1969 follow-up of a sample of the medical students surveyed by ASME in 1961 an attempt was made to compare the undergraduate record of trainee psychiatrists with doctors in other specialties. Interesting information on matters such as medical schools, work settings, and reading habits has emerged but the central question of the quality of psychiatrists remains unanswered. Further investigation, for example, medical students' attitudes to a career in psychiatry, is necessary as a preliminary to any recommendations to improve recruitment to the specialty.


"Gestation sac" volumes as determined by sonar in the first trimester of pregnancy. Sonar estimates of "gestation sac" volumes in the first trimester of pregnancy were made from a series of 319 measurements; the volumes estimated included the amniotic fluid, the extraembryonic coelom and the fetus. The technique employed involved the use of parallel section scans taken in series from one end of the gestation sac to the other, followed by planimetric measurements of the sac areas so produced. The potential errors which may be incurred in these measurements are discussed, and it is considered that the technique carries an overall possible error in the order of plus or minus 10 per cent. Growth curves produced from the measurements in this series show that the sac increases in volume from a mean of 1 ml. at 6 weeks to a mean of 100 ml. at 13 weeks, initially in an exponential fashion but latterly in a more linear manner. The mean values of the sonar gestation sac fluid volumes (after subtraction of the estimated fetal volume) in the 10 to 13 week range, show good correlation with those amniotic fluid values reported in the literature where direct measurements were made at the time of hysterotomy. As a method of assessing the maturity of a pregnancy this technique is of lesser value than the sonar measurement of fetal crown-rump length because of the relatively wider scatter of results. It has, however, found a useful place in clinical practice in the early sonar diagnosis of blighted ova or anembryonic pregnancies.


Ultrasonic  "B" scanning as a pregnancy test after less than six weeks amenorrhoea. Sixty patients with less than six weeks amenorrhoea were examined by ultrasonic "B" scanning for the detection of pregnancy. The results were compared with those obtained by histological examination of the aspirate after the patient had undergone menstrual regulation by vacuum aspiration of the uterine contents. The overall accuracy was 79 per cent with 15 per cent false positives. The accuracy after 39 days amenorrhoea was 95 per cent. The technique was compared with an immunological pregnancy test.


Continuous pethidine/diazepam infunsion during labour and its effects on the newborn. This paper presents a study comparing the effects on the mother and baby of a continuous intravenous infusion of pethidine and diazepam and of intramuscular pethidine and promazine. It is concluded that the administration of intravenous pethidine and diazepam is safe in labour provided that diazepam has not been given during the previous week.


Morphological changes in maternal lymphocytes in pregnancy. By use of staining with toluidine blue, the changes in the nucleoli of lymphocytes in 150 healthy women were studied. The patients were divided into five groups: non-pregnant women; women in the first, second and third trimesters of pregnancy; and women who were five days post partum. A progressive and statistically significant rise in the number of lymphocytes with micronucleoli was found during pregnancy and there was a fall after delivery. This increase was associated with a decrease in the incidence of ring-shaped nucleoli. There were no  significant changes in the absolute number of lymphocytes and in "active" lymphocytes during pregnancy.


Metabolism of prostaglandin F2alpha within the human uterus in early pregnancy. The potential for intra-uterine metabolism of prostaglandins in early pregnancy was studied by incubation of prostaglandin F2a (PGF2a) with tissues obtained by termination hysterectomy at 7 and 16 weeks gestation. Enzymes regulating the degradation of prostaglandins were demonstrated in placenta and membranes and to a lesser extent, in myometrium and decidua. It is believed that these enzymes may have a physiological role in maintaining the continuity of pregnancy, and that the high degradation of PGF2a in placenta and membranes may be one of the reasons that, for termination of pregnancy, far higher doses are required intra-amniotically than extra-amniotically.


The relationship of a "short period" to conception. The case records were examined of 918 patients delivered in 1963 at a London hospital. Comparisons were made between 52 patients who had a "short period" following the presumed date of conception and 155 randomly selected control patients. Patients who had a "short period" showed a significantly higher rate for the induction of labour (p smaller than 0-025) and a lower infant birthweight (0-1 smaller than p smaller than 0-2). Calculation of the menstruation-delivery interval using the "short period" rather than the last full period, gave a closer approximation to the duration of pregnancy found in the random control series of 155 patients.


Carbohydrate metabolism in the menstrual cycle. Blood sugar, serum insulin and growth hormone levels in response to 100 g. of oral glucose were studied under carefully controlled conditions on days 9, 18 and 27 of the menstrual cycle in 20 healthy women aged 18 to 20 years. The results indicate that blood sugar and serum insulin levels in response to oral glucose loading do not alter significantly in the menstrual cycle. Seven of the women had repeat studies on the same days three cycles later. No significant change in blood sugar or serum insulin levels occurred between cycles. The fasting growth hormone level was highest at the time of impending menstruation in 11 subjects (55 per cent) but the difference in mean values for the group as a whole was not statistically significant. It is concluded that the changes in endogenous hormone levels in a menstrual cycle do not produce significant alterations in carbohydrate metabolism in normal women in this age group. The reported divergence of blood glucose and insulin levels after oral glucose in the menstrual cycle may reflect the interaction of other factors such as age and parity on glucose tolerance. It is suggested that investigations into the metabolic effects of oral contraceptive steroids on these parameters should be performed in nulliparous women in a similar age range.


Maternal cardiomyopathy of pregnancy causing stillbirth. Cardiomyopathy of pregnancy developed in a healthy primigravida at 34 weeks gestation and was followed two weeks later by the spontaneous delivery of a fresh stillbirth. An immunological basis underlying both the mother's condition and the fetal death was suggested by the presence of myocardial antibodies in maternal and cord blood. Reference is made to two other cases in which a similar pathological process may have occurred.


The uterine cervix from adolescence to the menopause. During adolescence the physiological transformation zone of the cervix in the virgin undergoes limited change when compared to that of girls who are sexually promiscuous; the latter often show large areas of metaplastic squamous epithelium and the development of an atypical transformation zone. During pregnancy, particularly the first, the transformation zone increases in size and acquires increased amounts of squamous metaplastic epithelium, probably due to the effect of acidic vaginal fluid on endocervical columnar epithelium. Evidence exists for the development of an atypical transformation zone during pregnancy with its own variable neoplastic potential. After delivery the atypical epithelium of the cervix may be partly or completely removed with resulting regression of the lesion. Alternatively, neoplastic potential may alter or the lesion may remain static or recur in its previous form. After the childbearing era the physiological transformation zone undergoes minimal change until the menopause when the whole cervix shrinks. As the studies described above dramatically show, the sexually promiscuous woman at all ages has an increased risk of developing cervical neoplasia; this risk seems to be the highest in the third decade of life.


Neonatal hyperbilirubinaemia following the use of oxytocin in labour. A prospective study of 1353 labours and the relevant newborn failed to reveal any significant difference between the incidence of neonatal hyperbilirubinaemia (defined as a level of 12 mg. or more per 100 ml.) following spontaneous labour, and after labour induced or accelerated by oxytocin. The incidence of unexplained neonatal hyperbilirubinaemia after spontaneous labour was 6-3 per cent. Following induced labour however there was a highly significant (P less than 0-001) association between the mean total dose of oxytocin used for induction and the incidence of neonatal hyperbilirubinaemia. The proportion of babies who developed hyperbilirubinaemia increased in direct relation to the total dose of oxytocin used for the induction. In this series the incidence of hyperbilirubinaemia increased sharply when the total dose of oxytocin exceeded 20 units as it did hyperbilirubinaemia and birthweight, or duration of spontaneous labour. When labour was induced, however, the proportion on newborn babies with hyperbilirubinaemia increased with the duration of labour. The significance of these findings is discussed.


Serum cystine aminopeptidase and the normal weight baby in normotensive and hypertensive pregnancy. The levels of serum cystine aminopeptidase have been studied serially in normotensive and hypertensive mothers who delivered healthy, normal weight-for-dates babies. A normal range, both for enzyme level and maximum and minimum rates of increase of the enzyme during gestation, was established for each group, those of the hypertensive patients exhibiting different characteristics from the curves in normotensive patients. It is suggested that such differences be considered when assessing pregnancies complicated by hypertension.


Pregnancy in patients with coeliac disease. Successful pregnancy can occur both before and after the diagnosis and treatment of gluten-sensitive enteropathy. Abortion and intrauterine growth retardation are the main associated obstetric problems but their incidence may be reduced in the more severe cases by adherence to a strict gluten-free diet and dietary supplements. The present report concerns 60 pregnancies in 25 patients with coeliac disease.


Serum lipids in pregnancy and socio-economic status. Serum total and free cholesterol, phospholipids and erythrocyte free cholesterol levels were determined at four-weekly intervals from the first trimester until term in pregnant Nigerian women from three different socio-economic levels. The results showed a similar pattern of serum lipids in all three groups with a similar percentage increase at the peak levels. Serum phospholipids and total cholesterol levels were not significantly increased after the second trimester in contrast to report of a progressive hyperlipaemia in pregnant women of Western countries. Erythrocyte free cholesterol levels showed a mirror image of those of serum free cholesterol.


The prognostic value of human placental lactogen (HPL) levels in threatened abortion. The value of a single HPL estimation in serum was assessed in 337 patients with a threatened abortion between 7 and 27 weeks gestation. Serial assays were subsequently performed on 75 of these. A scheme is proposed whereby an HPL result can be given a "favourable", "equivocal" or "unfavourable" prognosis according to its level. Using this scheme a correct prognosis was obtained in 86 per cent of cases between 9 and 19 weeks gestation while an incorrect prognosis was obtained in 3 per cent of cases. There appeared to be little prognostic value after 19 weeks gestation though prior to 9 weeks the results were sufficiently promising to suggest that a more sensitive assay would be useful.


A review of 700 hysterotomies.

A review of 700 hysterotomies. We studied 700 hysterotomies carried out in Aberdeen between 1968 and 1972. The morbidity and mortality indicate that the operation is now outdated as a method of terminating pregnancy. A study of referral practice showed that there were many unneccessary delays.


Electroencephalographic changes after intra-amniotic prostaglandin F2alpha and hypertonic saline. Six major convulsions have been seen among 555 patients who had therapeutic second trimester abortion by intra-amniotic prostaglandin F2alpha (PGF2alpha). Five of 21 patients receiving intra-amniotic hypertonic saline and 5 of 16 patients receiving PGF2alpha showed abnormalities in electroencephalograms after treatment. These changes were relatively minor in all saline patients and in three receiving PGF2alpha, but two patients receiving PGF2alpha showed epileptiform spike activity after infusion. There was no correlation between the electroencephalographic (EEG) changes and peripheral venous levels of PGF2alpha or its metabolite, 13, 14 dihydro-15 keto PGF2alpha (15 keto-h2-f2alpha).


Operative culdoscopy.

Operative culdoscopy. A study of 257 operative culdoscopies done without general anaesthesia is presented and the advantages and limitations of the technique are detailed.


The investigation of ovarian function by measurement of urinary oestrogen and pregnanediol excretion. Urinary oestrogen and pregnanediol excretion was measued daily ("daily monitoring") for a complete cycle in 20 normally menstruating women, in one patient with an anovulatory cycle and for 28 days in a patient with secondary amenorrhoea. The measurements were also performed on urine specimens collected at weekly intervals for 4 to 6 weeks ("weekly tracking") from 506 patients with evidence of abnormal ovarian function. These included 9 patients with primary amenorrhoea, 132 patients with secondary amenorrheoa, 138 patients with oligomenorrhoea and 227 patients with evidence of ovarian dysfunction and cycle lengths of 25 to 42 days. The results were subjected to statistical analysis. In the normal cycles, ovulation could be identified on the criteria of a rising pregnanediol value reaching or exceeding 2-0 mg. per 24 hours for a period of 7 days or more. Valid conclusions on the overall mean oestrogen and pregnanediol values for a complete cycle could be made from the results of weekly tracking, irrespective of which day the tracking commenced. Correlations were obtained by comparing the mean and maximum urinary oestrogen values and the variability of the values with the evidence of ovarian function indicated by the clinical classifications of the patients, the duration of the disorders and the subsequent occurrence of uterine bleeding. Mean oestrogen values of 10 mug. per 24 hours or less were associated with lack of ovarian function. For values higher than this a discriminant function based on both the mean oestrogen value and the variability of the oestrogen values was useful in predicting onset of spontaneous menstruation. A single urine specimen collected 4 to 8 days before onset of menstruation showing a raised pregnanediol value of 2-0 mg. per 24 hours or more provided a valid test for ovulation in women with regular cycles, and a single urine specimen giving an oestrogen value of 10 mug per 24 hours or less gave a valid indication of absent ovarian function in women with amenorrhoea for two years or more. In all other circumstances serial sampling at weekly intervals provided a valid assessment of ovarian activity. Application of these principles allows the greatest amount of information on ovarian function to be obtained with the greatest economy of effort.


The resistant ovary syndrome.

The resistant ovary syndrome. A patient is described with primary amenorrhoea, raised FSH and LH levels and ovaries containing many primary follicles. These features were characteristic of the "resistant ovary syndrome". After ovarian wedge resection the patient began to menstruate.


Creatine phosphokinase (CPK) levels in pregnancy: a case report and a discussion of the value of CPK levels in the prediction of possible malignant hyperpyrexia. Creatine phosphokinase (CPK) levels during two consecutive pregnacies of a patient whose sister had died of malignant hyperpyrexia have been investigated. The results are compared with CPK levels of serum obtained from apparently normal people at similar stages of pregnancy. The rise of serum CPK occurring as a result of labour was much more pronounced in the malignant hyperpyrexia suspect.


Intracranial complications of transorbital stab wounds. Oscular and orbital injuries due to stab wounds may mask underlying serious intracranial damage. The correct clinical assessment and treatment of such cases require the attention of a team comprising a neurosurgeon, ophthalmologist, otolaryngologist, and plastic surgeon.


Euryblepharon.

Euryblepharon. The rare congenital anomaly of euryblepharon is reported in a male infant. It is suggested that euryblepharon is due to congenital hypoplasia or absence of the palpebral and lacrimal portions of the orbicularis oculi muscle.


Neurilemmoma of the oculomotor nerve.

Neurilemmoma of the oculomotor nerve. A case is presented of neurilemmoma of the left oculomotor nerve occurring in a 64-year-old hypertensive woman. The incipient tumour produced ptosis, limited inwards rotation of the eyeball, and persistent pupillary dilatation on the left side. The mechanism of this process is discussed, having regard to the interruption of the fibres innervating these structures by the tumour, and considering the mode of occurrence of the ocular and pupillary impairments. The patient also had generalized athero-sclerosis and fusiform aneurysm of the terminal part of the left internal carotid artery. The symptoms of headache, seizure, coma, and temporary right hemiparesis were considered as manifestations of hypertensive encephalopathy unrelated to the tumour.


The effect of growth hormone on calcium metabolism in the sheep. 1. The effect of subcutaneous administration of bovine growth hormone on calcium metabolism of nearly mature wether sheep has been studied by a combination of a radioactive technique and a nutrient balance technique. 2. Administration of growth hormone resulted in a significant increase in the rates of absorption of Ca, accretion of Ca into bone, resorption of Ca from bone and skeletal retention of Ca, and also in the sizes of the exchangeable Ca pools. 3. Retention of phosphorus was also significantly increased. 4. These changes suggest that the effect of growth hormone was to alter the Ca metabolism of nearly mature wethers to resemble that of younger, more actively growing animals. 5. Results are consistent with the theory that oestrogens may alter Ca metabolism of wethers by increasing growth hormone production.


Ascorbic acid and long-term anticonvulsant therapy in children. 1. Forty-eight children, who were long-term hospital patients, were investigated to assess their leucocyte ascorbic acid status. 2. Twenty-nine children had been receiving long-term anticonvulsants therapy, and these children did not have a significantly lower level of ascorbic acid than the untreated group, which did not have epilepsy. 3. The ascorbic levels of all subjects were low and seventeen had levels between 7-3 and 16-o mug/10-8 white blood cells.


Occurrence and conversion of anhydrolutein into dehydroretinol in a freshwater fish. 1. Lutein and anhydrolutein have been isolated from liver oils of freshly caught Saccobranchus fossilis. 2. S. fossilis initially contained more dehydroretinol than retinol; administration of anhydrolutein to the vitamin A-depleted fish resulted in the accumulation of dehydroretinol. 3. Anhydrolutein has also been isolated from the liver oils of lutein-treated, vitamin A-depleted fish. 4. The pathway of the conversion of lutein into dehydroretinol is discussed.


The absorption of iron, with or without supplements of single amino acids and of ascorbic acid, in healthy and Fe-deficient children. 1. Studies were done on the effect of ascorbic acid and five amino acids (histidine, cystine, cysteine, valine and glutamic acid) on intestinal iron absorption in a group of ninety Egyptian infants and young children, of which fifty-seven were healthy controls and thirty-three were suffering from Fe-deficiency anaemia. 2. Supplements tested promoted Fe absorption in healthy controls in the following order: valine larger than histidine larger than ascorbic acid. Cysteine, glutamic acid and cystine were found to have no significant effect. 3. Supplementation with valine, ascorbic acid and histidine also increased intestinal Fe absorption in anaemic subjects, but to a lesser extent than in controls. 4. Supplementation of haematinic therapy with these compounds is recommended. Their use is also suggested to improve the availability of the Fe content of everday diets.


An abnormality of the bone marrow associated with vitamin E deficiency in sheep. 1. Sheep fed on a maintenance ration of wheaten-hay chaff or of wheaten-hay chaff-lucerne-hay chaff (I : I, W/W) became deficient or incipiently deficient in vitamin E. 2. Degenerative changes were observed in bone marrow and muscle, and liver function was imparied in some animals. These abnormalities were not influenced by the vitamin B12 status of the animals or by a shortage of cobalt in the rumen. 3. Plasma ascorbic acid levels may not have been optimum, and folic may not have been fully utilized by some sheep. 4. Liver function responded faily rapidly to alpha-tocopheryl acetate, but skeletal muscle had not returned to normal after 28 weeks of treatment. A variable trend towards normal cellularity was found in bone marrow following supplementation with alpha-tocopheryl acetate. 5. A secondary deficiency or, alternatively, inefficient excretion or metabolism of a toxic material, may occur in vitamin E deficiency as a result of degenerative changes in the absorptive or excretory areas of the intestinal tract and be responsible for the bone marrow abnormality.


Effect of different regimens of early malnutrition on behavioural development and adult avoidance learning in Swiss white mice. 1. The effects of perinatal malnutrition on behavioural development and adult shuttle-box avoidance performance were studied in Swiss white mice. 2. Mice were malnourished (a) from the 7th day of gestation until birth, (b) from birth until weaning, or (c) during both gestation and the sucking period. 3. Pups born of protein-restricted mothers had reduced birth weights, retarded development and poor adult avoidance performance, even if reared from birth by well-nourished mothers. 4. Postnatal malnutrition, induced either by restricting maternal diet or by rearing in large litters, retarded development during the second half of lactation and lowered subsequent adult avoidance performance.


Effects of ingestion of organic selenium in adapted and non-adapted rats. 1. Rats were given moderate-selenium (4-5 mg/kg) or low-Se (0-5 mg/kg) diets during gestation and lactation. Their young were given diets with high (10 mg/kg), moderate or low Se contents from weaning, and groups of rats were killed at intervals during the 14-week experimental peroid. 2. Compared with young rats which received the low-Se diet, those which received the moderate- or high-Se diets had a high incidence of liver lesions and there were changes in liver Se content, haemoglobin concentration, packed cell volume, prothrombin activity, fibrinogen content, spleen weight, body water and serum glutamic-oxaloacetic and glutamic-pyruvic transaminas (L-aspartate : 2-oxoglutarate aminotransferase; EC 2.6.1.1 and L-alanine : 2-oxoglutarate aminotransferase; EC 2.6.1.2 respectively) and alkaline phosphatase (EC 3.1.3.1) activities. In those rats which received the high-Se diet the changes were more pronounced than in those which received the moderate-Se diet. 3. In young rats from dams given moderate-Se diets, which were themselves given the moderate-Se diet, the liver Se content decreased continuously, whereas rats given the same diet but from dams which had received the low-Se diet, the liver Se content increased continuously. There was a slight improvement of symptoms of Se toxicity in all groups by the 5th week of the experimental peroid. 4. The results suggest that there was an adaptation to chronic Se intake.


Effects of live weight and energy intake on nitrogen balance and total N requirement of lambs. 1. Results of 298 nitrogen balance studies from experiments with male cross-bred lambs, ranging in weight from 3 to 38 kg, which had been either fasted, or fed entirely on liqid diets of varying protein content at various energy intakes up to ad lib. intake, were used to quantitatively describe the effects of the amount and quality of absorbed protein, energy intake and live weight on N balance and total N requirement of lambs. 2. When N intake was less than the amount required, N balance was independent of energy intake, but linearly related to absorbed N and metabolic body-weight (live weight-O-75). In the fitted relationship, the coefficient of absorbed N was shown to be an estimate of the biological value of absorbed protein and the coefficient of metabolic body-weight was an estimate of the loss of endogenous N in both urine and faeces. For the milk-based diets used in the experiment biological value was 0-72 and the total endogenous N loss in urine and faeces was 148 mg N/kg per d. 3. When N intake was in excess of the amount required, N balance in lambs of a constant live weight increased linearly with metabolizable energy (ME) intake, at a rate that decreased with increasing live weight. Similarly at constant ME intake, N balance was a curvilinear decreasing function of metabolic body-weight, it was constant for lambs of all weights when ME intake was about 0-23 MJ/kg-0-75 per d, but it decreased linearly with increasing metabolic body-weight for ME intakes above this level. 4. N balance of fasted lambs was several times less than predicted by either of the relationships established for fed animals, and was found to be linearly related to metabolic body-weight. 5. The effects of energy intake and live weight on the total N requirement of lambs were determined. When total N requirement was expressed per unit of energy intake, it was found to be constant at 0-9 g N/MJ ME for all lambs irrespective of live weight when ME intake was 0-23 MJ/kg-0-75 per d. However, as ME intake/unit metabolic body-weight was raised above this level, N requirement/unit ME intake increased for lambs weighing less than c. 23 kg, but decreased for heavier animals.


Vitamin B12 nutrition and metabolism in the baboon (Papio cynocephalus). 1. Measurement of the vitamin B12 content of baboon tissues showed that the liver contained the highest concentration, followed by the pituitary, kidney, heart, spleen and pancreas. 2. The dietary vitamin B12 requirement of the baboon for the maintenance of satisfactory body stores was between 1 and 2 mug/d. 3. Satisfactory liver vitamin B12 stores were invariably associated with serum levels above 125 pg/ml, whereas liver levels were usually low when the serum level was below 50 pg/ml. 4. Increased methylmalonic acid (MMA) excretion after a valine load occurred when the liver vitamin B12 level was less than 0-40 mug/g. L- and DL-valine were approximately equally effective as precursors of MMA, whereas sodium propionate, whether given orally or intraperitoneally, was less effective. 5. The distribution of radioactivity along the wall of the intestinal tract after an oral dose of [57-Co]cyanocobalamin suggested that the distal half of the small intestine was the main site of vitamin B12 absorption. However, the utilization of vitamin B12 put direct into the middle part of the small intestine was much lower than that of an oral dose. 6. The unsaturated vitamin B12-binding capacity of baboon serum was not related to the serum vitamin B12 level. There was a significant difference between the unsaturated vitamin B12-binding capacities of the two subspecies of baboon (Papio cynocephalus cynocephalus and P. cynocephalus anubis) studied.


Metabolism of propionate in the tissues of the sheep gut. 1. The extent of propionate metabolism during absorption from the gut and the amounts of L-lactate formed and glucose utilized by the portal-drained viscera were determined in conscious sheep from measurements of portal venous blood flow and portal venous and aortic metabolite concentrations. The sheep were fasted overnight and given primed continuous intraruminal infusions of volatile fatty acids (VFA) at two rates, supplying propionate at 40.0 and 79.9 mmol/h. Measurements were made during the 5th and 6th hours of the infusion, when rumen liquor VFA concentrations were constant. 2. The rate of L-lactate formation by the portal-drained viscera was not affected by the VFA infusions and accounted for approximately 15% of the probably total lactate entry rate. 3. Considerable amounts of glucose were taken up by the portal-drained viscera, amounting to approximately 35% of the probable glucose entry rate. If this glucose was metabolized through the glycolytic pathway, this would at all times have accounted for the amounts of L-lactate formed. 4. Portal venous blood flow was positively correlated with VFA infusion rates and with the net amount of propionate appearing in the portal blood. 5. It is concluded that although propionate may be metabolized by the rumen epithelium, the unique pathway of L-lactate formation from propionate is of limited quantitative significance to the animal, although it may be of importance to the rumen epithelium itself.


The influence of essential fatty acids and food restriction on the specific activities of hepatic lipogenic and glutamate-metabolizing enzymes in the laying hen. 1. Extended feeding of an essential fatty acid (EFA)-deficient diet to laying hens increased liver size and liver lipid concentration. 2. The specific activities of hepatic lipogenic and glutamate-metabolizing enzymes were increased by feeding the EFA-deficient diet. 3. Restriction of daily food intake to 75% of ad lib. intake did not affect the response to dietary fat concentration. 4. Hepatic enzyme activities in hens fed on restricted amounts of food and killed just before, or after, the normal daily feeding time indicated no reduced capacity for lipogenesis at the earlier time.


Productive and unproductive lysozyme-chitosaccharide complexes. Equilibrium measurements. A method to determine both productive and unproductive lysozyme-chitosaccharide complexes has previously not been available. The method described in this paper uses a dye, Biebrich Scarlet, which forms a 1:1 complex with only part of the substrate binding site. Complex formation perturbs the spectrum of the compound and thus its dissociation constant can be determined (K-D equals 0.13 mM). The dissociation constants for three major enzyme-chitooligosaccharide complexes have also been determined: (1) chitooligosaccharides that bind only to sites A-C of lysozyme perturb the spectrum of the Biebrich Scarlet-lysozyme complex, without affecting the dissociation constant of the dye (K-u equals 0.01 mM); (2) chitooligosaccharides that interact with sites D-F displace the dye (K-S' equals 5-15 mM); (3) chitohexose forms a complex which involves the whole binding site and, therefore, also displaces Biebrich Scarlet. This complex, with a dissociation constant K-S equals 0.03 mM, is considered to be the productive one. The binding mechanism proposed on the basis of the results in this paper differs significantly from those considered previously.


Molecular dynamics and structure of the random coil and helical states of the collagen peptide, alpha 1-CB2, as determined by 13C magnetic resonance. Carbon-13 chemical shifts, spin-lattice (T1) and spin-spin (T2) relaxation times, and 13C-[1H] nuclear Overhauser enhancements (NOE) have been determined for the random coil and triple helical states of the alpha 1-CB2 fragment of rat skin collagen. Assignment of all aliphatic resonances of this 36 residue peptide in the random coil state (30 degrees) has been achieved with the aid of model polypeptides containing pyrrolidine residues. The chemical shifts and intensities of the Pro and Hyp C-gamma resonances show that (see article) 90% of the X-Pro and X-Hyp bonds are trans in both helix and coil conformations. From T1 measurements rotational correlation times (tau-eff) of ca. 0.45 nsec are calculated for interior C-alpha carbons in the coil, while taueff values of the side chain and near terminal carbons are found to be 2-9 times smaller. These results along with the narrow natural line widths (3-5 Hz) and maximal NOE values (2.8 plus or minus 0.3) demonstrate the high degree of backbone mobility, due to segmental motion, in the unordered state of the peptide. By contrast, the broad lines (50-90 Hz) and small NOE values (1.3 plus or minus 0.3) for the alpha carbons in the helical state (2 degrees) suggest much slower motion. The line widths and NOE values together with the C-alpha T1 values (0.025-0.040 sec) correspond to correlation times which are in reasonable agreement with those calculated for an axially symmetric rigid ellipsoid, undergoing rotational diffusion, having dimensions approximating those of a collagen-type triple helical aggregate of three alpha 1-CB2 chains. A satisfactory computer simulation of the experimental 2 degrees spectrum is obtained by assigning the narrow aliphatic resonances in the spectrum (line widths 5-40 Hz) to (a) carbons in the small amounts of alpha 1-CB2 (3 mol %) and alpha 1-CB1 (2.5 mol %) random coil conformations, (b) carbons in the flexible terminal triplets of the helix, and (c) Ala, Leu, and Phe methyl and phenyl carbons. The side chain carbon line widths obtained from the simulation--when compared with side chain line widths calculated for a rotating rigid ellipsoid with internal motion--indicate rapid axial reorientation of methyl and phenyl groups. With the exception of the Hyp residue the line widths suggest local motion for at least some carbons in most other side chain moieties. The Hyp C-beta and C-gamma line widths indicate the presence of little if any rapid Hyp ring motion.


Reconstitution of chromatin: mode of reassociation of chromosomal proteins. The mode of reassociation of Ehrlich ascites histones and non-histone proteins during chromatin reconstitution was studied by sodium dodecyl sulfate polyacryl-amide gel electrophoresis. In the procedure of Bekhor et al. (I. Bekhor, G. M. Kung, and J. Bonner, (1969), J. Mol. Biol. 39, 351) most of histones and non-histone proteins reassociate with DNA in the last dialysis step of the dissociated chromatin, that is the dialysis of the chromatin in 0.4 M NaCl-5 M urea against a dilute buffer. The reassociation of histones and non-histone proteins with DNA is more gradual in the procedure of L. Kleiman and R.-C. C. Huang [(1972), J. Mol. Biol. 64, 1]. However, in both procedures the bulk of the Ehrlich ascites non-histone proteins reassociate with DNA after the binding of histones to DNA. There are small amounts of non-histone proteins which reassociate with DNA before and at the same time as histones reassociate with DNA.


Chemical modifications of functional residues of fd gene 5 DNA-binding protein. The binding of gene 5 protein from bacteriophage fd to poly[d(A-T)], fdDNA, and poly(A) is accompanied by a dramatic reversal in the signs of the large ellipticity bands of the nucleic acid chromophores from 250 to 290 nm. The change in the circular dichroism of the DNA induced by the protein, which reaches a maximum at a protein to nucleotide molar ratio of 1:4, has been used as an assay of the alterations in binding of gene 5 protein to DNA accompanying changes in the ionic environment and subsequent to chemical modification of the protein. Divalent cations completely dissociate the gene 5 protein-fd DNA complex at 0.1 M, while 0.5 M monovalent cations are required. All cations are more effective in dissociating the complex with poly[d(A-T)] commensurate with the accompanying stabilization of the double helix to which gene 5 protein does not bind. Acetylation of all six lysyl residues and three of the five tyrosyl residues of the protein with N-acetylimidazole prevents complex formation. Removal of the three tyrosyl O-acetyl groups with hydroxylamine does not restore the binding of gene 5 protein to DNA. Tetranitromethane nitrates the same three tyrosyl residues (Tyr-26, Tyr-41, and Tyr-56 as determined by peptide mapping) and reduces the binding affinity of the protein for fd DNA by similar to 100-fold. The 19F NMR spectrum of gene 5 protein labeled with m-fluorotyrosine shows three surface and two buried fluorotyrosyl residues. All tyrosyl residues are protected from nitration in the complex with fd DNA, but acetylimidazole acetylates surface lysyl residues in the complex and dissociates it. The intrinsic circular dichroism of the acetylated and nitrated gene 5 proteins is not significantly altered. In contrast maleic anhydride reacts with the seven amino groups of the protein and changes the secondary structure to one similar to that present in 6 M guanidine-HCl. The single SH group of the native protein does not react with Ellman's reagent, but it reacts rapidly with one Hg2+ ion which unfolds the protein; fd DNA prevents reaction with Hg2+. Electrostatic forces may be as important as hydrogen bonding in maintaining the native structure of this protein. The lysyl groups of the protein, exposed in both the free protein and the DNA complex, appear to be of prime importance in DNA binding, probably through electrostatic interactions with the DNA binding, probably through electrostatic interactions with the DNA phosphate groups. Three tyrosyl residues also contribute to binding affinity through hydrogen bonding or intercalation. A model of gene 5 protein structure in relation to interactions with a tetranucleotide is presented.


The Bombyx mori silk proteins: characterization of large polypeptides. Proteins taken directly from the Bombyx mori silk gland have been separated and identified as either fibroin or sericin on the basis of their location within the gland and their amino acid composition. Molecular weights of these polypeptides have been determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and by agarose-guanidine chromatography. Fibroin consists of approximately equimolar amounts of two large (350,000) polypeptide chains. These may be the products of distinct fibroin alleles present in hybrid silkworm strains. Sericin, on the other hand, is composed of at least the three largest polypeptides (130,000-220,000) present in a mixture of proteins ranging in size from about 20,000 to 220,000.


Isolation, characterization, and amino acid sequence of melanotropins from camel pituitary glands. The isolation of two beta-melanotropins and two alpha-melanotropins from camel pituitary glands has been described, and their amino acid sequences have been determined. Two of them are identified as alpha-melanotropin and deacetylated alpha-melanotropin. There are also two beta-melanotropins whose structures are identical with the bovine hormone except that one has glycine in position 2 and the other glycine in position 2 as well as glutamine in position 8. The melanocyte-stimulating and lipolytic activities of these four camel melanotropins have been investigated by in vitro assay procedures.


Acetylation of nascent polypeptide chains on rat liver polyribosomes in vivo and in vitro. Rat liver polyribosomes may be acetylated in vivo utilizing [3-H]acetate as precursor and in vitro with [14-C]acetyl-CoA. The in vitro acetylation occurs almost completely in the amino terminal position while the in vivo acetylation (after correction for isotopic exchange and incorporation of tritium into nonacetyl positions of amino acids) was distributed equally between the amino terminal groups of a number of amino acids and the epsilon-amino groups of internal lysine residues. At least 50% of the labeled acetyl groups introduced in vivo as well as in vitro could be removed from polysomes as puromycin polypeptides or -peptidyl-tRNA. The acetylated polypeptides have been resolved by gel filtration into two components, one with an average molecular weight of 20,000 and the other of 4000-7000. The results presented indicate that the N-terminal acetylation of nascent growing polypeptides is a post initiation event that occurs on small peptides (40-70 amino acid residues) and depends on the presence of a polysome-bound acetyltransferase which differs from other cytoplasmic acetyltransferases which catalyze predominately the acetylation of internal amino groups of proteins.


Circular dichroism changes in galactosyltransferase upon substrate binding;. Circular dichroism studies with the galactosyltransferase isolated from bovine skim milk are described. Addition of UDP-galactose to the galactosyltransferase-Mn-2+ complex causes a decrease in the negative mean residue ellipticity in the 205-220-nm range and positive increases in the 265- and 275-290-nm ellipticity. These date are consistent with the view that a conformation change involving aromatic amino acid residues occurs upon the binding of UDP-galactose to the galactosyltransferase-Mn-2+ complex. No effects in the near-ultraviolet circular dichroism spectrum were observed upon the addition of UDP or glucose to the galactosyltransferase-Mn-2+ complex.


Purification and characterization of adrenodoxin reductase from bovine adrenal cortex. NADPH-adrenodoxin reductase from steer adrenal cortex mitochrondria has been purified to homogeneity (on sodium dodecyl sulfate polyacrylamide gel electrophoresis) by chromatography on DEAE-cellulose, Sephadex, and hydroxylapatite. A molecular weight of 51,500 was determined from sodium dodecyl sulfate polyacrylamide gel electrophoresis, while sedimentation equilibrium ultracentrifugation gave a value of 49,500. All of the flavine present was identified as FAD; 1 mol/52,000 g of protein. The reductase contained 1.7% carbohydrate (using glucose as standard) by weight. Homogeneous adrenodoxin reductase exhibited a typical oxidized flavoprotein absorbance spectrum, with maxima at 270, 376, and 450 nm, and gave an absorbance ratio A450/A270 of 0.122-0.128 (depending on the preparation). Reduction of the flavoprotein with NADPH or dithionite gave progressive bleaching of the 450-nm peak. The reductase was absolutely required, in the presence of adrenodoxin, for electron transfer from NADPH to cytochrome c or to particulate cytochrome P450. Adrenodoxin refuctase is obligatory for reconstitution of 11beta-hydroxylation activity using deoxycorticosterone as substrate, and for the side-chain cleavage of 20alpha-hydroxycholesterol or cholesterol. The specific activity of the homogeneous preparation in cytochrome c reduction is at least 17,000 nmol min-1 mg of protein-1, corresponding to a turnover number of 850 min-1. No evidence for the existence of multiple forms or subunits was obtained.


A pyrimidine nucleoside monophosphate kinase from rat liver. A pyrimidine nucleoside monophosphate kinase has been purified 2100-fold from rat liver. With ATP and dATP as phosphate donors the kinase uses CMP, dCMP, and UMP as phosphate acceptors. Ara-CMP is also phosphorylated by the kinase. In contrast to dCMP and UMP, CMP can be phosphorylated by dCTP. CTP and ara-CTP cannot substitute for dCTP. The stringent specificity of the phosphate donor site for ATP and dATP is lost when CMP serves as acceptor. All nucleoside triphosphates act as donors to a significant extent. No evidence has been found to suggest more than one enzyme. All activities, to different degrees, are strictly dependent upon preincubation at 37 degrees with a sulfhydryl reducing agent. Various reagents (85 mM) are ranked in order of increasing effectiveness of reactivation as follows: dithiothretiol greater than glutathione larger than or equal to 2-mercaptoethanol greater than L-cysteine greater than DL-alpha-lipoic acid. A NADP+-dependent thioredoxin (17 muM)-thioredoxin reductase system from Novikoff ascites rat tumor was found to be the most powerful reducing agent tested. CTP, dCTP, UTP, and dTTP (1 mM) do not affect the kinase activity regardless of the phosphate acceptor.


Chloride flux in bilayer membranes: the electrically silent chloride flux in semispherical bilayers. High resistance semispherical bilayer membranes of areas as large as 0.3 cm-2 were formed from a decane solution of synthetic diphytanoylphosphatidylcholine. These bilayers had a specific resistance of about 10-9 omega cm-2 and a specific capacitance of 0.38 mu F cm- minus 2 at 20 degrees in 0.1 M KCL. Under these conditions, chloride permeability was 6.8 times 10- minus 8 cm/sec. This electrically silen 36-Cl flux was found to be about 10-3-fold larger than the chloride current calculated from the electrical parameters of the system. The chloride flux in the bilayer was independent of the applied electrical field and was unaltered by addition of reducing agents to the ambient aqueous solutions. It was, however, substantially reduced when NO3 minus was substituted for Cl minus on the side of the bilayer initially free of 36-Cl, or if I minus was added to the aquesous phases in the concentration range of 0.001-0.1 M. These results strongly suggested that the electrically silent flux of 36-Cl is primarily a carrier mediated diffusion process in which phosphatidylcholine acts as the carrier species.


Laser Raman spectroscopy as a mechanistic probe of the phosphate transfer from adenosine triphosphate in a model system. Laser Raman spectroscopy has been used to study a phosphate transfer reaction from ATP to Pi or arsenate in dimethyl sulfoxide. The spectra support a mechanism involving Mg-2+ binding to the alpha or beta phosphates of ATP leaving the third phosphate free for the transfer reaction. The data also indicate the formation of a relatively stable intermediate which is facilitated by the presence of dimethyl sulfoxide and a dicarboxylic acid (maleate). The intermediate has a Raman spectrum with a band at 1090.5 cm- minus 1 similar to the end product ADP, but is formed much more rapidly. Since the model reaction has many features in common (e.g., activation by maleate) with the transfer reactions catalyzed by coupling factors from spinach chloroplast, Raman spectroscopy may also prove to be a useful tool in the elucidation of biological energy transfer reactions.


5-methyl-5,6,7,8-tetrahydropteroyl oligo-gamma-L-glutamates: synthesis and kinetic studies with methionine synthetase from bovine brain. The synthesis of 5-methyl-5,6,7,8-tetrahydropteroly tri-, pena-, and heptaglutamate has been accomplished by reductive methylation of the tetrahydropteroyl oligoglutamate with formaldehyde, followed by purification on DEAE-Sephadex. The corresponding [5-14-C]methyltetrahydropteroyl oligoglutamates were prepared from 14-CH-2-0, and tested as substrates for methionine synthetase (EC 2.1.1.13) ISOLATED FROM BOVINE BRAIN. In all cases, the polyglutamate conjugates were better substrates (lower Km, higher Vmax) than the corresponding monoglutamate forms. In addition, the nonradioactive methyltetrahydropteroyl oligoglutamates inhibited the methylation of homocysteine by methyltetrahydrofolate. This indicates that the monoglutamate and polyglutamates compete for the same enzyme, and established a role for the ubiquitous methyltetrahydropteroyl oligoglutamates in mammalian methionine biosynthesis.


A light scattering investigation of the propylurea dissociation of human hemoglobin A. The subunit dissociation of human hemoglobin A by propylurea in several liganded and chemically modified states was investigated by light scattering molecular weight methods. The dissociation data were analyzed by means of the equation developed in our earlier studies: deltaF degrees D EQUALS TO DELTAF degrees D, W - 2N'RTKB[D], where deltaF degrees D and deltaf degreees D,W represent the free energy of dissociation of hemoglobin tetramers into half-molecules consisting of alpha beta dimers in the presence and in the absence of propylurea, KB is the binding constant of the urea to the average peptide unit, [D] is its concentration, and N' is the number of amino acid sites exposed per half-molecule on dissociation. It is found that the dissociation of oxyhemoglobin, cyanmethemoglobin, and N-ethylmaleimide oxyhemoglobin is characterized by essentially the same N' value of 15 to 21 plus or minus 3, that are close to the 19 amino acid residues per surface which comprise the smaller alpha beta contact area, seen in the X-ray crystallographic model of horse hemoglobin of Perutz and coworkers. Due to the very low degree of dissociation of deoxyhemoglobin, only a very approximate estimate of N' of about the same order of magnitude could be obtained for this form of the protein. In contrast, a significantly lower value of N' was obtained with bis(maleimidomethyl) ether modified oxyhemoglobin of 8 plus or minus 3, that is cross-linked at cysteine residue F9 (93)beta and histidine residue FG4 (97) beta in the same beta chains. Our results suggest that alterations caused by the presence of the cross-linking reagent reflect both the loss in amino acid residues that can interact with the urea at the blocked segments of the polypeptide chains in the dissociated state of hemoglobin and the changes in accessibility of some of the amino acid residues perturbed by the introduction of the reagent in the parent tetrameric form.


Purification of an apparent rat liver prothrombin precursor: characterization and comparison to normal rat prothrombin. Current evidence would suggest that prothrombin is synthesized from a liver precursor molecule in a vitamin K dependent step which involves the attachment of calcium binding groups to the precursor. A protein has now been isolated from the liver of warfarin-treated rats which has the properties predicted for this precursor. The purified precursor is a glycoprotein with a molecular weight indistinguishable from rat prothrombin. Both electrophoretic and isofocusing analyses indicate that the precursor is less negatively charged than prothrombin. Specific proteolysis of the precursor by thrombin, taipan snake venom, or clotting factor Xa yielded fragments indistinguishable from those formed by similar proteolysis of prothrombin. The rate of activation of the precursor to thrombin by factor Xa and Ca-2+ was not stimulated by the addition of phospholipid, while prothrombin activation is greatly stimulated under these conditions. All of the data obtained are consistent with the hypothesis that the protein isolated is a precursor to prothrombin, and that under the influence of vitamin K, this precursor is converted to prothrombin by the addition of a number of acidic calcium binding groups.


Isolation of peptides from the carboxyl carrier subunit of transcarboxylase. Role of the non-biotinyl peptide in assembly. Transcarboxylase is made up of a central hexameric subunit (S20,W similar 12 S), three peripheral dimeric metallo subunits (S20,W similar to 5 S), and six biotinyl carboxyl carrier subunits (S20,W similar to 1.3 S). The results presented here show that the carboxyl carrier subunit is required for assembly of the 12S and 5S subunits into the oligomer. However, only a portion of the subunit is required for assembly. On treatment of transcarboxylase briefly with trypsin at pH 6.3 extremely susceptible peptide bonds of the carboxyl carrier protein are cleaved releasing biotinyl peptides of about similar to 66 and similar to 40 residues. The resulting trypsinized transcarboxylase, though enzymatically inactive, remains essentially intact as judged by its hydrodynamic and molecular sieving properties. The modified enzyme can be dissociated at pH 8 to the central 12S subunit and peripheral 5S subunit to which the residual portion(s) of the cleaved carboxyl carrier protein is still attached. These components can then be separated by molecular sieving. The residual portion of the carboxyl carrier protein (non-biotinyl peptide) can then be isolated by dissociation of the 5S subunit complex at pH 9 and by chromatography over Bio-Gel A-1.5m. The isolated non-biotinyl peptide has been shown to contain the combining domain of the 1.3SE carboxyl carrier protein since it causes combination of the 12S and 5S subunits. Active enzyme is formed by combination of the intact carboxyl carrier protein and the 12S and 5S subunits and an inactive oligomer of similar size is formed if the non-biotinyl peptide is used in place of the carboxyl carrier protein. The similar to 66- and similar to 40-residue biotinyl peptides, which are released by the trypsin treatment, apparently occur on an exposed portion of the enzyme. This portion of the carboxyl carrier protein apparently serves to place the biotinyl group adjacent to the two substrate sites of the enzyme, one of which is on the peripheral subunit and the other on the central subunit. Thus the carboxyl carrier protein has two functions: one portion holds the 12S and 5S subunits in juxtaposition and the other portion orients the biotinyl group adjacent to the substrate sites so that it may function as a carboxyl carrier between the sites.


Evidence that the two partial reactions of transcarboxylation are catalyzed by two dissimilar subunits of transcarboxylase. The results presented here show that isolated subunits of transcarboxylase specifically catalyze the two partial reactions of transcarboxylation as shown in eq 1-3. The 12S central subunit is active in the transcarboxylation with methylmalonyl-CoA but inactive with oxalacetate and the peripheral metallo 5S subunit is active in the transcarboxylation with oxalacetate but inactive with methylmalonyl-CoA. These subunits, likewise, are specific for the reverse partial reactions; the central subunit catalyzing transfer from the carboxylated biotinyl group to propionyl-CoA to yield methylmalonyl-CoA and the peripheral subunit to pyruvate to yield oxalacetate. Thus, the central subunit contains the sites for the CoA esters (methylmalonyl-CoA and propionyl-CoA) and the peripheral metallo subunits for the keto acids (oxalacetate and pyruvate). In the overall reaction the biotinyl carboxyl carrier protein acts as a shuttle to carry the carboxyl groups between the two subunits. Biotin and certain biotin analogs are inactive in these partial reactions but the similar to 40- or similar to 66-residue biotinyl peptides, which are derived from the carboxyl carrier protein, are active. Transcarboxylase can be reconstituted from its isolated subunits and a comparison was made of the rate of the overall reaction when the subunits were assembled, as in the intact enzyme, with that obtained when the reaction was catalyzed by the nonassembled subunits. In the latter case, since the biotinyl carboxyl carrier subunit must diffuse from one subunit to the other, the overall reaction is much slower than with the assembled subunits. The reaction with trypsinized transcarboxylase from which the similar to 66-residue and similar to 40-residue biotinyl peptides have been stripped, likewise, was slow even though the biotinyl peptides were added to the reconstitution mixture. The 12SH and 5SE subunits remain assembled after trypsin treatment but the biotinyl peptides apparently do not combine firmly or properly with the trypsinized enzyme and the biotinyl group apparently must oscillate as a carboxyl carrier between the two sites on the subunits by diffusion.


Partial purification and properties of calf thymus deoxyribonucleic acid dependent RNA polymerase III. DNA-dependent RNA polymerase III (nucleosidetriphosphate: RNA nucleotidyltransferase, EC 2.7.-7.6) has been isolated and partially purified from calf thymus tissue. Significant amounts of enzyme III are present in this tissue (up to 15% of the total activity of thymus homogenates). This enzyme has been characterized with respect to its chromatographic properties, broad ammonium sulfate optimum (0.04-0.2 M), template requirements, divalent metal optima, and its unique alpha-amanitin sensitivity (50% inhibition of activity occurring at an alpha-amanitin concentration of 10 mug/ml).


DNA ligase activity in chromatin and its analogs. Rejoining of DNA strands in polylysine-DNA complexes and in reconstituted chromatins. A highly purified DNA ligase from rat liver nuclei has been tested on DNA containing single-strand breaks ("nicks"); the DNA was present in several types of complexes which were chosen to serve as models for chromatin. These model systems included complexes of polylysine or histones with DNA as well as reconstituted chromatin preparations. In all these cases, the limit of ligase sealing was measured as a function of the ratio of polypeptide or protein to DNA. With an excess of either polylysine or histones, the ligase is totally prevented from sealing nicks in the DNA. However, at ratios of histones to DNA similar to those occurring in chromatin, about half of the nicks are accessible to the ligase. In the reconstitution of chromatin, the proteins are dissociated from the DNA by exposure to high ionic strength either with or without urea. If such procedures are carried out in the presence of labeled nicked DNA, the proteins will redistribute over this ligase substrate as well. When the chromatin is reconstituted at protein/DNA ratios similar to those occurring in chromatin, once more only about half of the nicks are accessible to the ligase. Similar results were obtained with preparations reconstituted with either rat liver or duck reticulocyte chromatin. The rate of ligase action has been measured on a variety of the complexes. While the rate falls as the DNA is increasingly covered with polylysine or histones, this is largely or entirely due to the decrease in concentration of sealable sites. At saturating concentrations of these DNA complexes, the original rate on uncovered DNA is approached.


The contribution of RNA and non-histone proteins to the circular dichroism spectrum of chromatin. This paper is an investigation of the contribution of low salt extractable RNA and non-histone proteins to the circular dichroism of chromatin. Circular dichroism (CD) of chromatin above 250 nm is due mainly to DNA and is different from that of DNA free in solution. In addition, to a smaller extent, we find that low salt extractable RNA and/or non-histone protein side chain chromophores contribute significantly to the spectra in this region and account for the major differences observed among the CD spectra of chromatins isolated from the five tissues studied; pig cerebellum, myeloma, calf thymus, chick embryo brain, and chick erythrocytes.


Synthesis of rhodopsin and opsin in vitro. Isolated bovine retinas have been used to study the synthesis of rhodopsin and the renewal of photoreceptor rod outer segments. Both leucine and glucosamine served as radioactive tracers to follow rhodopsin synthesis. In both cases the rod outer segment preparations contained large amounts of labeled macromolecules chromatographically distinct from rhodopsin, the latter representing only about 10% of the high molecular weight labeled material. However, electrophoresis on polyacrylamide gels with sodium dodecyl sulfate indicated that as much as 60-68% of the radioactivity coincided with opsin, the apoprotein of rhodopsin. Treatment of labeled rod outer segments with 9-cis-retinal caused much of the non-rhodopsin label to be converted to isorhodopsin. After such treatment the fraction of the label in visual pigment rose from about 10 to 51% with leucine as the radioactive tracer and to 78% with glucosamine. Similar treatment of bleached outer segments labeled with leucine gave identical results with complete regeneration of isorhodopsin (lambda max 487 nm) which then accounted for 56% of the labeled macromolecules. No such conversion occurred in controls lacking 9-cis-retinal. Both 9-cis- and 11-cis-retinal were effective but all-trans-retinal was ineffective in producing the conversion. Under in vitro conditions opsin appears to be accumulated in the outer segment prior to the addition of retinal.


The F3-F2a1 complex as a unit in the self-assembly of nucleoproteins. A specific and stable interaction between histones f3 and f2a1 was demonstrated to take place in the absence of DNA. When a mixture of these histones was subjected to velocity sedimentation under conditions in which the separate histones are aggregated and precipitate, the mixture of f3 and f2a1 remained soluble and these histones appeared to cotransport through the gradient, indicating the establishment of an isolatable, stable f3-f2a1 complex. This isolated complex subsequently binds to DNA quantitatively to form nucleohistone. Stoichiometry data strongly suggest that histones f3 and f2a1 bind to DNA as a unit; this is the only type of f2a1 binding to DNA that can take place under mild conditions. Histone f1 can act as a modifier of the f3-f2a1-DNA interactions by augmenting the formation of the f3-f2a1 complex and consequently enhancing the overall binding of these histones to DNA. No significant interactions of histones f2b and f2a2 with other histones could be demonstrated. Because of the findings reported here and the known affinity characteristics of the arginine-rich histones to DNA in native chromatin (in particular their stimultaneous extraction from chromatin by salt), we suggest that the (f3 + f2a1)-DNA complex is a structural component of native chromatin. We would also like to propose that, in vivo, histones may possess a considerable amount of quaternary structure, which would greatly increase the specificity of their role as potential regulators of the structure and function of the eucaryotic chromosomes.


Iodohexestrols. I. Synthesis and photoreactivity of iodinated hexestrol derivatives. The five possible ortho-iodinated hexestrols, containing from 1-4 iodines per molecule, have been synthesized as potential iodine-containing estrogenic ligands. After chromatographic separation and purification, each derivative has been fully characterized by spectroscopic methods. [3-H]-3-Iodohexestrol, [131-I]- and [125-I]-3-iodohexestrol, and [125-IA1-3,5-diiodohexestrol have also been prepared. Photolysis of 3,5-diiodohexestrol in methanol results in rapid deiodination to 3-iodohexestrol; further reduction to hexestrol is slower. Photolysis of 3-iodohexestrol in benzene gives 3-phenylhexestrol.


Fusion of fatty acid containing lecithin vesicles. The rate and temperature at which fusion of purified homogeneous lecithin vesicles containing several per cent fatty acid occurs have been determined by analysis of proton nuclear magnetic resonance (nmr) spectra. Dimyristoyllecithin with myristic acid as 2% of the lipid was found to fuse rapidly at temperatures between 17 and 20 degrees, and dimyristoyllecithin with 4% lauric acid was found to fuse rapidly at temperatures between 11 and 15 degrees, while dimyristoyllecithin with 4% palmitic acid did not fuse at an appreciable rate anywhere in the range 17-37 degrees. These results, along with data on dipalmitoyl- and dilauroyllecithin, are discussed in terms of a possible dependence on separation of fatty acid rich phase especially conducive to fusion.


Isolation of messenger-like ribonucleoproteins. Subribosomal and polyribosomal messenger ribonucleoproteins (mRNPs) were isolated from Ehrlich ascites tumor cells by a method involving sedimentation of polyribosomal and subribosomal particles, dissociation with EDTA, and rate-zonal sedimentation. The fractions containing mRNA protein particles were applied to glass fiber filters and extensively washed with buffer containing 0.5 M KCl. The eluted material was demonstrated to be an RNA-protein complex containing poly(A)-rich RNA, heterogeneous in size, and free of 18S or 28S rRNA. mRNA function for the RNA was suggested by its ability to direct protein synthesis in a cell-free protein-synthesizing system derived from wheat germ embryos. Analysis of the proteins associated with subribosomal and polyribosomal mRNPs by iodination and sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed at least seven similar proteins. The apparent molecular weights of the three most prominent proteins were 78,000, 52,000, and 34,000. Analysis of reticulocyte polyribosomal mRNPs revealed an increased prominence of the 78,000 and 52,000 molecular weight proteins relative to the other protein bands.


Solubilization of the Semliki Forest virus membrane with sodium dodecyl sulfate. The dissociation of Semliki Forest virus induced by increasing concentrations of the anionic detergent sodium dodecyl sulfate was studied using density gradient centrifugation. Detectable binding to the virus started well below the critical micellar concentration of the detergent and increased thereafter with increased detergent concentration. At 4 degrees there were about 11,000 binding sites per virus particle with an average association constant of about 10-5 M-1. The extent of virus dissociation could be controlled both by the detergent concentration and by the temperature. At 4 degrees only disruption ("lysis") of the virus membrane could be observed. At 20 degrees most of the membrane was solubilized into lipoprotein complexes, and the nucleocapsid dissociated into RNA and protein. Complete delipidation of the viral membrane proteins was achieved at 30 degrees at a detergent concentration still below the critical micellar concentration.


A reinvestigation of the amino-terminal sequence of human parathyroid hormone. The sequence of the amino-terminal portion of human parathyroid hormone, particularly the identity of residues 22, 28, and 30 (the subject of discrepancies in recent published reports), has been reexamined by two basic methods of structural analysis. A fresh lot of human parathyroid hormone isolated from pooled adenoma tissue was analyzed by Edman degradation with identification of critical residues by thin-layer chromatography and gas-liquid chromatography. In the second approach, -14C or tritiated amino acids were incorporated during biosynthesis of the human hormone in slices of parathyroid glands in vitro; the appropriate amino acid residues were then determined as the -14C or tritiated phenythiohydantoin derivatives of the amino acid after Edman degradation, or by peptide isolation after appropriate cleavage with endopeptidase, or both. The results confirm our previous findings that residue 22 is glutamic acid, residue 28 is leucine, and residue 30 is aspartic acid.


Kinetic model of the protein-mediated phosphatidylcholine exchange between single bilayer liposomes. The phosphatidylcholine exchange protein from beef liver catalyzes the exchange of phosphatidylcholine between single bilayer liposomes (Hellings et al. (1974), Eur. J. Biochem. 47, 601). A model has been proposed which describes the kinetics of this exchange. Steady-state equations have been derived from the model and have been used for the derivation of the theoretical rate equation. Computer analysis shows a good fit with the experimental results. It follows from the analysis that the apparent dissociation constant of the exchange protein-liposome complex decreases with an increasing phosphatidic acid content of the liposomes. This suggests that in this model system it is the phospholipid composition of the membranes involved that regulates the amount of exchange protein available to function as a carrier of phosphatidylcholine.


Covalent structure of collagen: amino acid sequence of alpha1-CB3 of chick skin collagen. The amino acid sequence of alpha1-CB3, a peptide containing 149 residues obtained from the central portion of the alpha1(I) chain of chick skin collagen by cyanogen bromide cleavage, has been determined. As in the other sequences from the helical region of collagen chains, the repeating triplet Gly-X-Y extends throughout the length of the peptide. These data allow a comparison of the sequence to that of alpha1-CB3 from calf and rat skin collagens. As compared with calf, the chick peptide contains 14 amino acid differences, whereas it contains 17 residue differences from the rat peptide. Thus, the sequence identity level is 91 and 89 percent, respectively, in comparison to the calf and rat peptides. These values are significantly greater than the value of 97 percent observed between the peptide of the two mammalian species and reflect the greater phylogenetic distance of the species compared.


Covalent structure of collagen: amino acid sequence of alpha1-CB6A of chick skin collagen. The amino acid sequence of chick skin collagen alpha1-CB6A, a peptide containing 107 residues obtained from the helical region near the carboxy-terminus of the alpha1(I) chain by cyanogen bromide cleavage, has been determined. This was accomplished by automated Edman degradation of the hydroxylamine-produced fragments and of the tryptic peptides prepared with and without prior maleylation. The data show that this portion of the alpha1(I) chain from chick skin is identical in 90 percent of the residues to the corresponding peptide region of calf skin collagen reported previously.