Exploiting the high-resolution crystal structure of Staphylococcus aureus MenH to gain insight into enzyme activity

Alice Dawson, Paul K. Fyfe, Florian Gillet, William N. Hunter

    Research output: Contribution to journalArticle

    11 Citations (Scopus)

    Abstract

    Background: MenH (2-succinyl-6-hydroxy-2,4-cyclohexadiene-1-carboxylate synthase) is a key enzyme in the biosynthesis of menaquinone, catalyzing an unusual 2,5-elimination of pyruvate from 2-succinyl-5-enolpyruvyl-6-hydroxy-3-cyclohexadiene-1-carboxylate.

    Results: The crystal structure of Staphylococcus aureus MenH has been determined at 2 angstrom resolution. In the absence of a complex to inform on aspects of specificity a model of the enzyme-substrate complex has been used in conjunction with previously published kinetic analyses, site-directed mutagenesis studies and comparisons with orthologues to investigate the structure and reactivity of MenH.

    Conclusions: The overall basic active site displays pronounced hydrophobic character on one side and these properties complement those of the substrate. A complex network of hydrogen bonds involving well-ordered water molecules serves to position key residues participating in the recognition of substrate and subsequent catalysis. We propose a proton shuttle mechanism, reliant on a catalytic triad consisting of Ser89, Asp216 and His243. The reaction is initiated by proton abstraction from the substrate by an activated Ser89. The propensity to form a conjugated system provides the driving force for pyruvate elimination. During the elimination, a methylene group is converted to a methyl and we judge it likely that His243 provides a proton, previously acquired from Ser89 for that reduction. A conformational change of the protonated His243 may be encouraged by the presence of an anionic intermediate in the active site.

    Original languageEnglish
    Article number19
    Pages (from-to)-
    Number of pages11
    JournalBMC Structural Biology
    Volume11
    DOIs
    Publication statusPublished - 22 Apr 2011

    Keywords

    • ALPHA/BETA-HYDROLASE FOLD
    • MENAQUINONE VITAMIN-K2 BIOSYNTHESIS
    • ESCHERICHIA-COLI
    • CATALYTIC MECHANISM
    • SEQUENCE ALIGNMENT
    • SYNTHASE
    • IDENTIFICATION
    • SPECIFICITY
    • REACTIVITY
    • COMPLEX

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