title: Structures of Helicobacter pylori shikimate kinase reveal a selective inhibitor-induced-fit mechanism.
abstract: Shikimate kinase (SK), which catalyzes the specific phosphorylation of the 3-hydroxyl group of shikimic acid in the presence of ATP, is the enzyme in the fifth step of the shikimate pathway for biosynthesis of aromatic amino acids. This pathway is present in bacteria, fungi, and plants but absent in mammals and therefore represents an attractive target pathway for the development of new antimicrobial agents, herbicides, and antiparasitic agents. Here we investigated the detailed structure-activity relationship of SK from Helicobacter pylori (HpSK). Site-directed mutagenesis and isothermal titration calorimetry studies revealed critical conserved residues (D33, F48, R57, R116, and R132) that interact with shikimate and are therefore involved in catalysis. Crystal structures of HpSK·SO(4), R57A, and HpSK•shikimate-3-phosphate • ADP show a characteristic three-layer architecture and a conformationally elastic region consisting of F48, R57, R116, and R132, occupied by shikimate. The structure of the inhibitor complex, E114A • 162535, was also determined, which revealed a dramatic shift in the elastic LID region and resulted in conformational locking into a distinctive form. These results reveal considerable insight into the active-site chemistry of SKs and a selective inhibitor-induced-fit mechanism.
authors.affiliation: Institute of Molecular and Cellular Biology and Department of Life Sciences, National Tsing Hua University, Hsinchu, Taiwan.
authors.forename: Wen-Chi
authors.initials: WC
authors.lastname: Cheng
authors.forename: Yen-Fu
authors.initials: YF
authors.lastname: Chen
authors.forename: Hung-Jung
authors.initials: HJ
authors.lastname: Wang
authors.forename: Kai-Cheng
authors.initials: KC
authors.lastname: Hsu
authors.forename: Shuang-Chih
authors.initials: SC
authors.lastname: Lin
authors.forename: Tzu-Jung
authors.initials: TJ
authors.lastname: Chen
authors.forename: Jinn-Moon
authors.initials: JM
authors.lastname: Yang
authors.forename: Wen-Ching
authors.initials: WC
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002151:Calorimetry; D020134:Catalytic Domain / Q000235:genetics; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D055808:Drug Discovery; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005798:Genes, Bacterial; D016480:Helicobacter pylori / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012765:Shikimic Acid / Q000378:metabolism; D055672:Static Electricity
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D012765:Shikimic Acid; D017853:Phosphotransferases (Alcohol Group Acceptor); C022642:shikimate kinase