title: Identification and characterization of D-xylose reductase involved in pentose catabolism of the zygomycetous fungus Rhizomucor pusillus.
abstract: Rhizomucor pusillus NBRC 4578 efficiently produces ethanol from lignocellulosic biomass because of its ability to ferment not only d-glucose, but also d-xylose. When the strain was cultivated on d-xylose, ethanol was gradually formed in the culture medium with a decrease in d-xylose and the simultaneous accumulation of xylitol, which suggested that the strain catabolized d-xylose with d-xylose reductase (XR) and xylitol dehydrogenase (XDH). XR (RpXR) was purified to homogeneity from the crude extract prepared from the mycelia of the strain grown on d-xylose. The purified enzyme was found to be NADPH-dependent and prefer pentoses such as d-xylose, d-ribose, and l-arabinose as substrates. Isolation of the genomic DNA and cDNA of the xyl1 gene encoding RpXR revealed that the gene was interrupted by two introns and the exon of the gene encoded a protein composed of 322 amino acids with a Mr of 36,724. Phylogenetic analysis showed that RpXR is more related to 4-dihydromethyltrisporate dehydrogenases from Mucoraseae fungi rather than the previously reported fungal XRs. Quantitative real-time PCR indicated that transcription of the xyl1 gene was marked in the presence of d-xylose and l-arabinose, but was week in the presence of d-glucose. These biochemical and expression analyses suggest that RpXR is involved in the catabolism of l-arabinose as well as d-xylose. This is the first report of the purification, characterization, and gene cloning of XR from zygomycetous fungi.
authors.affiliation: Biotechnology Research Center and Department of Biotechnology, Toyama Prefectural University, 5180 Kurokawa, Imizu, Toyama 939-0398, Japan. Electronic address: hkomeda@pu-toyama.ac.jp.
authors.forename: Hidenobu
authors.initials: H
authors.lastname: Komeda
authors.affiliation: Biotechnology Research Center and Department of Biotechnology, Toyama Prefectural University, 5180 Kurokawa, Imizu, Toyama 939-0398, Japan.
authors.forename: Shino
authors.initials: S
authors.lastname: Yamasaki-Yashiki
authors.affiliation: Graduate School of Science and Engineering, University of Toyama, 3190 Gofuku, Toyama 930-8555, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Hoshino
authors.affiliation: Biotechnology Research Center and Department of Biotechnology, Toyama Prefectural University, 5180 Kurokawa, Imizu, Toyama 939-0398, Japan.
authors.forename: Yasuhisa
authors.initials: Y
authors.lastname: Asano
mesh_terms: D000449:Aldehyde Reductase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D001089:Arabinose / Q000378:metabolism; D003001:Cloning, Molecular; D050542:D-Xylulose Reductase / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D000431:Ethanol / Q000378:metabolism; D019143:Evolution, Molecular; D005947:Glucose / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D010429:Pentoses / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D020103:Rhizomucor / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D012266:Ribose / Q000235:genetics; D014993:Xylitol / Q000378:metabolism; D014994:Xylose / Q000378:metabolism
chemical_list: D010429:Pentoses; D000431:Ethanol; D009249:NADP; D012266:Ribose; D014994:Xylose; D001089:Arabinose; D000449:Aldehyde Reductase; D050542:D-Xylulose Reductase; D005947:Glucose; D014993:Xylitol
keywords: 4-Dihydromethyltrisporate dehydrogenase; Pentose catabolism; Rhizomucor pusillus; Zygomycetous fungus; d-Xylose reductase