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Acta Cryst. (2014). A70, C443
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Dihydrodipicolinate synthase (DHDPS) is an enzyme found in most bacterial species and regulates the production of cell wall precursors necessary for the life of the organism. Specifically, DHDPS catalyzes the condensation of pyruvate and aspartate-β-semialdehyde (ASA) to produce dihydrodipicolinic acid leading to the synthesis of cell wall precursors lysine and mesodiaminopimelate. DHDPS is regulated through feedback inhibition when lysine binds at a location distinct from the reactive site. The mechanism by which lysine remotely disrupts catalysis is not well understood. A clear understanding of how the natural inhibitor, lysine, binds to DHDPS and what effects this has on the machinery of the enzyme will be invaluable for development of novel antibiotic leads. Analysis of DHDPS crystals with and without inhibitors has revealed structural changes that appear to link the allosteric and active sites. Our ongoing research examines the validity of observed structural changes in the mechanism of allosteric inhibition.

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Acta Cryst. (2014). A70, C477
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Campylobacter jejuni (Cjj) is the leading cause of gastroenteritis in humans, its glycoconjugates are known to include sugars in the furanose ring conformation. In particular, Cjj HS:41 strains are associated with the development of Guillian-Barré syndrome (GBS) and produce a key virulence factor of the bacterium, namely furanose-based capsular polysaccharides (CPS). To date, the enzymes responsible for furanose biosynthesis in Cjj HS:41 CPS are poorly understood. CPS sequencing has revealed three genes, annotated as glf1-3 via homology to known bacterial UDP-D-galactopyranose mutase (UGM), could be involved in the biosynthetic pathways. Our laboratories are interested in the structure-function relationship behind recognition and discrimination within such pyranose-furanose mutases. Enzymology studies have shown Glf1 is a flavoprotein responsible for isomerization between GDP-6d-D-altro-heptopyranose and GDP-6d-D-altro-heptofuranose, the latter being a major component of CPS. This GDP-altro-heptopyranose mutase (GaHM) activity is the first example of a heptose-recognizing mutase. Given that many of the UGM active site and FAD binding site residues are conserved in GaHM, the catalytic mechanism is likely similar to that of UGM. In order to establish key features of the enzyme, detailed structural information is required. We report here on the structure of Cjj GaHM, which has been co-crystallized with GDP, in both oxidized and reduced states. Due to low sequence identity with bacterial UGMs, Se-MET SAD phasing was ultimately employed to solve the structure. We will also discuss our recent crystallization efforts with Cjj GaHM in the presence of GDP-sugar substrate derivatives. Ultimately, the structural information gleaned from this study could lead to the identification of a new inhibitors targeting the CPS biosynthetic pathway.
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