Biological and Pharmaceutical Bulletin
Online ISSN : 1347-5215
Print ISSN : 0918-6158
ISSN-L : 0918-6158
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Cloning, Expression and Purification of Arylsulfate Sulfotransferase from Eubacterium A-44
Bomi KimYang-Jin HyunKeun-Sook LeeKyoichi KobashiDong-Hyun Kim
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JOURNAL FREE ACCESS

2007 Volume 30 Issue 1 Pages 11-14

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Abstract

A gene (astA) encoding arylsulfate sulfotransferase (ASST), which transfers a sulfate group from phenolic sulfate esters to phenolic acceptors, was cloned from a Eubacterium A-44 genomic library. The probe (1.5 kb fragment) for the astA gene was prepared from the PCR product of the primers produced using two internal amino acid sequences of ASST, which had been purified from Eubacterium A-44. The astA gene was cloned into the pKF3vector. Its sequence revealed a 1863 bp open reading frame (ORF) encoding a protein containing 620 amino acids with a secretary signal peptide, and showed 91% homology (identity) to Eubacterium rectale IIIH previously reported. The cloned astA gene was expressed under the T7 promoter of the expression vectors, pET-39b(+) and pET-26b(+), in Escherichia coli BL21 (DE3), and the expressed ASSTs were purified using His Bind column chromatography. The specific activities of the purified ASSTs were 25.6 μmol/min/mg and 37.1 μmol/min/mg, respectively.

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© 2007 The Pharmaceutical Society of Japan
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