Purification of branched-chain amino acid aminotransferase from Helicobacter pylori NCTC 11637 |
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Authors: | M. Saito K. Nishimura S. Wakabayashi T. Kurihara Y. Nagata |
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Affiliation: | (1) Department of Materials and Applied Chemistry, College of Science and Technology, Nihon University, Tokyo, Japan;(2) Department of Applied Chemistry, Junior College, Nihon University, Funabashi, Chiba;(3) Department of Life Sciences, Graduate School of Life Science, University of Hyogo, Kamigohri, Hyogo, Japan;(4) Institute for Chemical Research, Kyoto University, Uji, Kyoto, Japan |
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Abstract: | Branched-chain amino acid aminotransferase was purified by several column chromatographies from Helicobacter pylori NCTC 11637, and the N-terminal amino acid sequence was analyzed. The enzyme gene was sequenced based on a putative branched-chain amino acid aminotransferase gene, ilvE of H. pylori 26695, and the whole amino acid sequence was deduced from the nucleotide sequence. The enzyme existed in a homodimer with a calculated subunit molecular weight (MW) of 37,539 and an isoelectric point (pI) of 6.47. The enzyme showed high affinity to 2-oxoglutarate (K m = 0.085 mM) and L-isoleucine (K m = 0.34 mM), and V max was 27.3 μmol/min/mg. The best substrate was found to be L-isoleucine followed by L-leucine and L-valine. No activity was shown toward the D-enantiomers of these amino acids. The optimal pH and temperature were pH 8.0 and 37 °C, respectively. |
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Keywords: | : Branched-chain amino acid aminotransferase – Purification – Helicobacter pylori – ilvE gene |
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