title: Purification and characterization of membrane-associated hydrogenase from the deep-sea epsilonproteobacterium Hydrogenimonas thermophila.
abstract: Membrane-associated hydrogenase was purified from the chemolithoautotrophic epsilonproteobacterium Hydrogenimonas thermophila at 152-fold purity. The hydrogenase was found to be localized in the periplasmic space, and was easily solubilized with 0.1% Triton X-100 treatment. Hydrogen oxidation activity was 1,365 micromol H(2)/min/mg of protein at 80 degrees C at pH 9.0, with phenazine methosulphate as the electron acceptor. Hydrogen production activity was 900 micromol H(2)/min/mg of protein at 80 degrees C and pH 6.0, with reduced methyl viologen as the electron donor. The hydrogenase from this organism showed higher oxygen tolerance than those from other microorganisms showing hydrogen oxidation activity. The structural genes of this hydrogenase, which contains N-terminal amino acid sequences from both small and large subunits of purified hydrogenase, were successfully elucidated. The hydrogenase from H. thermophila was found to be phylogenetically related with H(2) uptake hydrogenases from pathogenic Epsilonproteobacteria.
authors.affiliation: Laboratory of Marine Microbiology, Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Kitashirakawaoiwake-cho, Sakyo-ku, Kyoto, Japan. h1ro5h1@hotmail.co.jp
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Nishimura
authors.forename: Yuki
authors.initials: Y
authors.lastname: Kitano
authors.forename: Takahiro
authors.initials: T
authors.lastname: Inoue
authors.forename: Keigo
authors.initials: K
authors.lastname: Nomura
authors.forename: Yoshihiko
authors.initials: Y
authors.lastname: Sako
mesh_terms: D000595:Amino Acid Sequence; D001709:Biotechnology; D002462:Cell Membrane / Q000201:enzymology*; D020565:Epsilonproteobacteria / Q000166:cytology* / Q000201:enzymology*; D006864:Hydrogenase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D021381:Protein Transport; D012995:Solubility; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D006864:Hydrogenase