Cloning and characterization of a bacterial cell-bound type B carboxylesterase from Bacillus sp. BP-7.
A clone producing halos on tributyrin plates was isolated from a genomic library of Bacillus sp. BP-7. The insert contained an open reading frame that coded for a protein of 487 amino acids with homology to carboxylesterases. The cloned enzyme showed clear preference for esters of short-chain fatty acids, being classified as an esterase. Maximum activity was found at 45 degrees C and pH 7.5. The enzyme displayed stability in the pH range from 6 to 9.5, and at temperatures from 4 degrees to 45 degrees C. Zymogram analysis of the protein revealed a molecular mass of 53 kDa and a pI of 5.1. The enzyme showed homology to members of the bacterial subclass of type B carboxylesterases, a set of proteins potentially useful for biotechnological applications.
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Department of Microbiology, Faculty of Biology, University of Barcelona, Av. Diagonal 645, 08028-Barcelona, Spain.
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D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D043182:Carboxylesterase; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D016680:Genome, Bacterial; D015698:Genomic Library; D006923:Hymecromone / Q000378:metabolism; D007525:Isoelectric Focusing; D008066:Lipolysis; D008969:Molecular Sequence Data; D009596:Nitrophenols / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity
D001426:Bacterial Proteins; D009596:Nitrophenols; D011994:Recombinant Proteins; D006923:Hymecromone; D002265:Carboxylic Ester Hydrolases; D043182:Carboxylesterase