Molecular Cloning and High-Level Expression in Escherichia coli of Fungal α-Galactosidase from Absidia corymbifera IFO 8084

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概要

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• A cDNA encoding the α-galactosidase of Absidia corymbifera IFO 8084 was cloned and sequenced. The cloned DNA has a single open-reading frame consisting of 2190 base pairs, and the deduced amino acid sequence revealed that the mature enzyme consisted of 730 amino acid residues with a molecular mass of 82, 712 Da. The native structure of the α-galactosidase of A.corymbifera IFO 8084 was determined to be a tetramer. Comparison with amino acid sequences of other α-galactosidase showed high homology with sequences of members of family 36. An expression vector, pET32Trx/galα, was constructed by introducing the cDNA coding region into a thioredoxin fusion system, pET32-Ek/LIC. The resulting transformant, pET32Trx/galα, overproduced the active enzyme as a thioredoxin fused form in the host Escherichia coli. By using His-binding metal affinity chromatography, recombinant α-galactosidase was purified to homogeneity in a single step. The purified recombinant fusion α-galactosidase showed properties very similar to the native α-galactosidase from A.corymbifera IFO 8084.

• 社団法人日本生物工学会の論文
• 2000-08-25

著者

• Yokota Atsushi

  Laboratory of Microbial Physiology, Research Faculty of Agriculture, Hokkaido University

• Asano Kozo

  Laboratory of Applied Microbiology, Graduate School of Agriculture, Hokkaido University

• TOMITA FUSAO

  Laboratory of Applied Microbiology, Graduate School of Agriculture, Hokkaido University

• SAITO KATSUICHI

  Laboratory of Applied Microbiology, Graduate School of Agriculture, Hokkaido University

• 横田 篤

  Laboratory Of Microbial Physiology Research Faculty Of Agriculture Hokkaido University

• Yokota Atsushi

  Lab. Of Microbial Physiology Res. Fac. Of Agriculture Hokkaido Univ. Kita 9 Nishi 9 Kita-ku Sapporo

• 横田 篤

  北大 大学院農学研究院

• Yokota Atsushi

  Laboratory Of Bioorganic Chemistry

• Yokota A

  Laboratory Of Applied Microbiology Department Of Bioscience And Chemistry Faculty Of Agriculture Hok

• Yokota Atsushi

  Hokkaido Univ. Sapporo Jpn

• Yokota Atsushi

  Laboratory Of Applied Microbiology Department Of Bioscience And Chemistry Faculty Of Agriculture Hok

• Asano Kozo

  Laboratory Of Applied Microbiology Graduate School Of Agriculture Hokkaido University

• Asano Kozo

  北海道大学農学部応用生命科学専攻

• Tomita Fusao

  Laboratory Of Applied Microbiology

• Tomita Fusao

  Department Of Agricultural Chemistry Faculty Of Agriculture Hokkaido University

• Saito Katsuichi

  Department Of Upland Agriculture National Agricultural Research Center For Hokkaido Region

• Saito Katsuichi

  Memuro Upland Farming Research Station National Agricultural Research Center For Hokkaido Region

• Saito Katsuichi

  Laboratory Of Applied Microbiology Graduate School Of Agriculture Hokkaido University

• Yokota A

  Hokkaido Univ. Hokkaido Jpn

• Saito K

  Department Of Upland Agriculture National Agricultural Research Center For Hokkaido Region

• BAIK SANG

  Laboratory of Applied Microbiology, Graduate School of Agriculture, Hokkaido University

• Baik S

  Laboratory Of Applied Microbiology Graduate School Of Agriculture Hokkaido University

• Baik Sang

  Laboratory Of Applied Microbiology Graduate School Of Agriculture Hokkaido University

• Asano Kozo

  Laboratory Of Applied Microbiology Graduate Faculty Of Agriculture Hokkaido University

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