Our long-range goal is to understand how the cell works in terms of its component organelles and membranes. Cellular function will be studied on several levels. D-beta-hydroxybutyrate dehydrogenase is a membrane-bound enzyme which has been purified in homogeneous form. It is a phospholipid-requiring enzyme, i.e., the enzyme is totally inactive without lecithin. A comprehensive study of this enzyme is planned to elucidate the molecular basis for the role of lipid. On the level of the membrane, we are engaged in a comprehensive study of sarcoplasmic reticulum from muscle which regulates contraction and relaxation via release and uptake of calcium. We have recently been successful in dissociating this membrane and reassembling it both with regard to calcium pumping as well as in morphology and composition. This breakthrough now makes possible the study of principles of membrane assembly, as well as the basis of calcium uptake and release. Both biochemical and biophysical techniques will be used to characterize the molecular association of phospholipids in serum lipoproteins and in the active D-beta-hydroxybutyrate dehydrogenase-lecithin complex, and in sarcoplasmic reticulum membranes. We will improve our technology for the prolonged storage of tissue and subsequent subcellular fractionation for potential use with clinical material. These techniques will be applied to the study of the intracellular movement of cholesterol and components important in intracellular regulation. Finally, focus of the studies will be on hormonal regulation important in diabetes, including glucagon and insulin in normal and diabetic rats.
NIH ExPorter
NIH ExPorter