Journal
JOURNAL OF MOLECULAR BIOLOGY
Volume 363, Issue 2, Pages 383-394Publisher
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.jmb.2006.07.057
Keywords
lipopolysaccharide; heptose; glycosyltransferase; crystal structure
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Lipopolysaccharides constitute the outer leaflet of the outer membrane of Gram-negative bacteria and are therefore essential for cell growth and viability. The heptosyltransferase WaaC is a glycosyltransferase (GT) involved in the synthesis of the inner core region of LPS. It catalyzes the addition of the first L-glycero-D-manno-heptose (heptose) molecule to one 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo) residue of the Kdo(2)-lipid A molecule. Heptose is an essential component of the LPS core domain; its absence results in a truncated lipopolysaccharide associated with the deep-rough phenotype causing a greater susceptibility to antibiotic and an attenuated virulence for pathogenic Gram-negative bacteria. Thus, WaaC represents a promising target in antibacterial drug design. Here, we report the structure of WaaC from the Escherichia coli pathogenic strain RS218 alone at 1.9 angstrom resolution, and in complex with either ADP or the non-cleavable analog ADP-2-deoxy-2-fluoro-heptose of the sugar donor at 2.4 angstrom resolution. WaaC adopts the GT-B fold in two domains, characteristic of one glycosyltransferase structural superfamily. The comparison of the three different structures shows that WaaC does not undergo a domain rotation, characteristic of the GT-B family, upon substrate binding, but allows the substrate analog and the reaction product to adopt remarkably distinct conformations inside the active site. In addition, both binary complexes offer a close view of the donor subsite and, together with results from site-directed mutagenesis studies, provide evidence for a model of the catalytic mechanism. (c) 2006 Published by Elsevier Ltd.
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