Grant: Discovery Grants Program - Individual

Our LONG-TERM GOALS are to elucidate the mechanism(s) of biosynthesis of lipopolysaccharides (LPS) and to utilise this information for designing novel enzyme inhibitors with antimicrobial properties. Our MODEL SYSTEM focuses mainly on the LPS molecule of Escherichia coli, but also the LPS of opportunistic bacteria such as Burkholderia cenocepacia and Pseudomonas aeruginosa. We are particularly interested in two areas of biological and biotechnological interest: (i) the biosynthesis of LPS core oligosaccharide, and (ii) the structure-function analysis of bacterial glycosyltransferases (GCTs).    (i) We will FOCUS the current research program on the identification and characterization of a novel pathway for the synthesis of the core oligosaccharide LPS precursor, ADP-L-glycero-D¬-manno-heptose (ADP-LDHep). This molecule is a building block required for the synthesis of the core oligosaccharide in most Gram-negative bacteria. Mutants with a heptoseless core display a dramatic increase in permeability to hydrophobic compounds and they survive very poorly in the host. Thus, targeting the enzymes involved in the ADP-LDHep biosynthesis pathway may result in the development of new antimicrobials.    (ii) In the second part of our proposal, we will continue with the detailed characterization of the four glycosyltransferases (GCTs) involved in the synthesis of the O7 antigen repeating subunit. We have described a novel analogue of the isoprenoid lipid undecaprenol phosphate, which facilitates the characterization of glycosylation products and used it to identify the function of WbbD, the first of the four GCTs for O7 antigen synthesis. We will to continue with the characterization of the remaining glysosyltransferases WbbC, WbbB, and WbbA, and initiate crystallization trails for each of these enzymes.    In terms of SIGNIFCANCE, the areas of research described in this proposal build on solid preliminary data from the PI's laboratory and his collaborators, and will contribute to further our basic knowledge on LPS biosynthesis, while at the same discover novel strategies to curtail infection.