Functional assignment of Glu386 and Arg388 in the active site of l-galactono-γ-lactone dehydrogenase

Nicole G.H. Leferink, Mac Donald F. Jose, Willy A.M. van den Berg, Willem J.H. van Berkel

    Research output: Contribution to journalArticlepeer-review

    Abstract

    The flavoenzyme l-galactono-γ-lactone dehydrogenase (GALDH) catalyzes the terminal step of vitamin C biosynthesis in plants. Little is known about the catalytic mechanism of GALDH and related aldonolactone oxidoreductases. Here we identified an essential Glu-Arg pair in the active site of GALDH from Arabidopsis thaliana. Glu386 and Arg388 variants show high Km values for l-galactono-1,4-lactone and low turnover rates. Arg388 is crucial for the stabilization of the anionic form of the reduced FAD cofactor. Glu386 is involved in productive substrate binding. The E386D variant has lost its specificity for l-galactono-1,4-lactone and shows the highest catalytic efficiency with l-gulono-1,4-lactone.

    Original languageEnglish
    Pages (from-to)3199-3203
    Number of pages5
    JournalFEBS Letters
    Volume583
    Issue number19
    DOIs
    Publication statusPublished - 6 Oct 2009

    Keywords

    • Flavoprotein
    • l-Galactono-1,4-lactone dehydrogenase (EC 1.3.2.3)
    • Site-directed mutagenesis
    • VAO family
    • Vitamin C

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