Analysis of WbaP, an enzyme involved in initiation of group 1 capsule assembly in Escherichia coli serotype K30
This thesis reports an investigation of WbaP, a UDP-Gal:phosphoryl-polyprenol Gal-1-phosphate transferase required for the first step in biosynthesis of capsular polysaccharide in 'E. coli' serotype K30. WbaP is a member of the p_olyisoprenyl-phosphate h_exose-1-p_hosphate t_ransferases (PHPT) that catalyze the transfer of hexose-1-P residues from an activated nucleotide diphosphosugar donor to a lipid carrier. These enzymes play an important role in the virulence of many bacteria as its members initiate biosynthesis of O antigens and capsular polysaccharides. Representatives of this family are integral membrane proteins with an N-terminal domain containing several transmembrane helices and a C-terminal domain with a cytoplasmic active site. Little is known about the structural and functional details of these enzymes. The experimental data reported here provides evidence to localize the catalytic domain of WbaP to the C-terminus. Strategies for overexpression of WbaP, determination of its membrane topology and characterization of its basic biochemical properties are described.