A novel nicotinoprotein aldehyde dehydrogenase involved in polyethylene glycol degradation

T. Ohta, A. Tani, K. Kimbara, F. Kawai

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    30 Citations (Scopus)

    Abstract

    A gene (pegC) encoding aldehyde dehydrogenase (ALDH) was located 3.4 kb upstream of a gene encoding polyethylene glycol (PEG) dehydrogenase (pegA) in Sphingomonas macrogoltabidus strain 103. ALDH was expressed in Escherichia coli and purified on a Ni-nitrilotriacetic acid agarose column. The recombinant enzyme was a homotetramer consisting of four 46.1-kDa subunits. The alignment of the putative amino acid sequence of the cloned enzyme showed high similarity with a group of NAD(P)-dependent ALDHs (identity 36-52%); NAD-binding domains (Rossmann fold and four glycine residues) and catalytic residues (Glu225 and Cys259) were well conserved. The cofactor, which was extracted from the purified enzyme, was tightly bound to the enzyme and identified as NADP. The enzyme contained 0.94 mol NADP per subunit. The enzyme was activated by Ca 2+, but by no other metals; no metal (Zn, Fe, Mg, or Mn) was detected in the purified recombinant enzyme. Activity was inhibited by p-chloromercuric benzoate, and heavy metals such as Hg, Cu, Pb and Cd, indicating that a cysteine residue is involved in the activity. Enzyme activity was independent of N,N-dimethyl-p-nitrosoaniline as an electron acceptor. Trans-4-(N,N- dimethylamino)-cinnamaldehyde was not oxidized as a substrate, but the compound worked as an inhibitor for the enzyme, as did pyrazole. The enzyme acted on n-aldehydes C2-C14) and PEG-aldehydes. Thus the enzyme was concluded to be a novel Ca2+-activating nicotinoprotein (NADP-containing) PEG-aldehyde dehydrogenase involved in the degradation of PEG in S. macrogoltabidus strain 103.

    Original languageEnglish
    Pages (from-to)639-646
    Number of pages8
    JournalApplied Microbiology and Biotechnology
    Volume68
    Issue number5
    DOIs
    Publication statusPublished - Sep 2005

    ASJC Scopus subject areas

    • Biotechnology
    • Applied Microbiology and Biotechnology

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