title: Investigation of the role of a conserved glycine motif in the Saccharomyces cerevisiae xylose reductase.
abstract: All yeast xylose reductases, with the exception of that from Schizosaccharomyces pombe, possess the catalytic and coenzyme-binding elements from both the aldo-keto reductase and short-chain dehydrogenase-reductase (SDR) enzyme families in their primary sequences. In the Saccharomyces cerevisiae xylose reductase (XR), the SDR-like coenzyme-binding GXXXGXG motif (Gly motif) is located between residues 128 and 134, with the third Gly residue being replaced by an Asp. We used site-directed mutagenesis to study the role of this SDR-like Gly motif in the S. cerevisiae xylose reductase. Site-directed mutagenesis of the individual conserved Gly residue positions (G128A, G132A, D134G, and D134A) did not significantly affect the specific activity, kinetic constants (K(m), K(cat), and K(cat)/K(m)), or dissociation constants (K(d)) in any of the variants compared with the wild type. Deletion of the entire Gly motif produced an unstable protein that could not be purified. These results indicate that the SDR-like Gly motif likely provides support to the overall structure of the enzyme, but it does not contribute directly to coenzyme binding in this XR.
authors.affiliation: Department of Environmental Biology, University of Guelph, Guelph, Ontario, Canada, N1G 2W1.
authors.forename: Byron C H
authors.initials: BC
authors.lastname: Chu
authors.forename: Hung
authors.initials: H
authors.lastname: Lee
mesh_terms: D000449:Aldehyde Reductase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020816:Amino Acid Motifs / Q000502:physiology*; D005998:Glycine / Q000737:chemistry*; D016297:Mutagenesis, Site-Directed; D009249:NADP / Q000378:metabolism; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D009249:NADP; D000449:Aldehyde Reductase; D005998:Glycine