A study of bacterial deoxysugars is proposed using two complementary experimental approaches: I. Detailed examination of individual enzymes involved in the biosynthesis of 6-deoxyhexoses: 1) Thymidinediphosphoglucose-4,6-hydrolase, the enzyme initiating 6-deoxyhexose biosynthesis will be studied to increase the understanding of structure and function for "half-site" enzymes. The problem will be explored by: a) chemical modifications of enzyme protein; b) studies concerning the process of enzyme-subunit association and disassociation; c) kinetic analysis with synthetic substrate and coenzyme analogues and studies on specific enzyme inhibitors. 2) Comparative studies will be undertaken on UDP-galactose-4-epimerase, another "half-site" enzyme with analogous properties to the enzyme studied in 1). 3) TDP-L-rhamnose dehydrogenase and TDP-6-deoxy-L-talose dehydrogenase. a) Purification of these enzymes by affinity chromatography; b) kinetic analysis with synthetic substrate and coenzyme analogues. II. In a synchronized bacterial culture, the level of TDPG-4,6-hydrolase will be correlated with the amount of L-rhamnose incorporated into the cell wall lipopolysaccharide. One objective of these studies will be to find compounds capable of interferring with deoxyhexose biosynthesis (part I of this project). These inhibitors will be tested for their ability to inhibit 6-deoxyhexose biosynthesis in the in vivo system just described.
NIH ExPorter
NIH ExPorter