Cloning of the pepX Gene of Lactobacillus helveticus IFO3809 Encoding Salt-Tolerant X-Prolyl Dipeptidyl Aminopeptidase and Characterization of the Enzyme
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概要
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X-prolyl dipeptidyl aminopeptidase (X-PDAP) from Lactobacillus helveticus IFO3809 expressed nearly full activity under high salt conditions, such as 2 M NaCl. We cloned and sequenced the pepX gene for X-PDAP. The calculated M_r of deduced X-PDAP (803 amino acids) was 90,847 and the protein was distantly related (35 to 44% identity) to known X-PDAPs of Lactobacillus sp. including L. helveticus CNRZ32 (40% identity). Native and recombinant X-PDAP were purified to homogeneity from both L. helveticus IFO3809 and Escherichia coli DH5α harboring the pepX gene on a plasmid, respectively. The native enzyme appeared to be a dimer of 220 kDa, as estimated by gel filtration column chromatography. It hydrolyzed an X-prolyl-linkage, but not prolyl- or X-prolyl-X-peptide bonds, and tolerated up to 2 M NaCl as well as some other chlorides of monovalent cations. Determination of the flanking sequences revealed two divergent genes. The upstream region of the pepX gene encodes oppA gene for a putative oligopeptide permease, while the downstream region encodes tnp gene specifying a possible transposase of the IS3 family. The oppA gene shares a 176 bp-promoter region with pepX in the intergenic region, implying a relationship between this oligopeptide transport system and X-PDAP.
- 社団法人日本生物工学会の論文
- 2002-06-20
著者
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Fujii Mikio
Foods Research Group Foods Division Asahi Chemical Industry Co. Ltd.
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Itoh Yoshifumi
Division Of Applied Microbiology National Food Research Institute
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KIMURA Keitarou
Division of Applied Microbiology, National Food Research Institute
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Fujii M
Japan Tobacco Inc. Shizuoka Jpn
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Itoh Y
Division Of Applied Microbiology National Food Research Institute
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NAGASAWA ATSUSHI
Foods Technology Center, Food Business Division, Japan Tobacco Inc.
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Nagasawa Atsushi
Foods Technology Center Food Business Division Japan Tobacco Inc.
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Fujii Mikio
Foods Technology Center Food Business Division Japan Tobacco Inc.
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Kimura Keitarou
Division Of Applied Microbiology National Food Research Institute
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