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Biosci Biotechnol Biochem. 1994 Jan;58(1):111-6.

Cloning of Pseudomonas fluorescens carboxylesterase gene and characterization of its product expressed in Escherichia coli.

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1
Department of Life Science, Korea Advanced Institute of Science and Technology, Kusung-dong Yusung-gu, Taejon.

Abstract

A gene (estC) coding for an esterase (esterase III) of Pseudomonas fluorescens was cloned into Escherichia coli JM83. DNA sequencing showed a single open reading frame with GTG as a translation initiation codon for esterase III. This was confirmed by N-terminal amino acid sequence analysis of the purified esterase III protein from an E. coli clone. The promoter sequence and a potential Shine-Dalgarno sequence were followed by the coding sequence of the estC gene. The amino acid sequence deduced from the nucleotide sequence contains the consensus active site sequence, G-X-S-X-G, of serine esterase. The esterase III expressed in an E. coli clone was purified by ion-exchange chromatography and gel filtration. The native form of the enzyme was a monomer with a molecular weight of 41,000. The results of substrate specificity and the inhibitor studies suggest that this enzyme is a carboxylesterase (EC 3.1.1.1) and a serine residue is present at the active site of the enzyme.

PMID:
7764506
DOI:
10.1271/bbb.58.111
[Indexed for MEDLINE]
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