Cloning and Sequence Analysis of the Gene for Glucodextranase from Arthrobacter globiformis T-3044 and Expression in Escherichia coli Cells
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概要
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The gld gene for glucodextranase from Arthrobacter globiformis T-3044 was cloned by using a combination of gene walking and probe methods and expressed on the recombinant plasmid pGD8,which was constructed with pUC118,in Escherichia coli cells. The enzyme gene consisted of a unique open reading frame of 3,153 bp. The comparison of the DNA sequence data with the N-terminal and 6 internal amino acid sequences of the purified enzyme secreted from A. globiformis T-3044 suggested the enzyme was translated from mRNA as a secretory precursor with a signal peptide of 28 amino acids residues. The deduced amino acids sequence of the mature enzyme contained 1,023 residues, resulting in a polypeptide with a molecular mass of 107,475 daltons. The deduced sequence showed about 38% identity to that of the glucoamylase from Clostridium sp. G0005. The glucodextranase activity of transformant harboring pGD8 was about 40 mU/ml at 30℃ for a 16-h culture. Although the GDase that was produced from the transformant was shorter than authentic GDase by 2 amino acid residues at the N-terminal end side, its enzymatic properties were almost same as the authentic one. Two kinds of genes, dex1 and dex2,for endo-dextranases from A. globiformis T-3044 were also cloned into Escherichia coli cells. The N-terminal of the purified endo-dextranase from A. globiformis T-3044 agreed with the deduced amino acid sequence, after the 33rd alanine residue, of only the dex1 gene for endo-dextranase. This result suggests that the endo-dextranase is translated from mRNA as a secretory precursor with a signal peptide of 32 amino acids residues. The deduced sequence of endo-dextranase 1 and endo-dextranase 2 showed about 93% and 65% identity with that of known endo-dextranase from Arthrobacter sp. CB-8,respectively.
- 社団法人日本農芸化学会の論文
- 1999-12-23
著者
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Oguma Tetsuya
Noda Institute For Scientific Research
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KOBAYASHI Mikihiko
National Food Research Institute
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Kitao Satoshi
Research And Development Division Kikkoman Corporation
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Tobe K
Kikkoman Corp. Chiba Jpn
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KUROKAWA Toshiko
Noda Institute for Scientific Research
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TOBE Kouichiro
Research and Development Division, Kikkoman Corporation
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Tobe Kouichiro
Research And Development Division Kikkoman Corporation
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Kobayashi Mikihiko
National Food Research Institute Ministry Of Agriculture Forestry And Fisheries
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