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A new esterase showing similarity to putative dienelactone hydrolase from a strict marine bacterium, Vibrio sp. GMD509.

Abstract
Vibrio sp. GMD509, a marine bacterium isolated from eggs of the sea hare, exhibited lipolytic activity on tributyrin (TBN) plate, and the gene representing lipolytic activity was cloned. As a result, an open reading frame (ORF) consisting of 1,017 bp (338 aa) was found, and the deduced amino acid sequence of the ORF showed low similarity (< 20%) to alpha/beta hydrolases such as dienelactone hydrolases and esterase/lipase with G-X(1)-S-X(2)-G sequence conserved. Phylogenetic analysis suggested that the protein belonged to a new family of esterase/lipase together with various hypothetical proteins. The enzyme was overexpressed in Escherichia coli and purified to homogeneity. The purified enzyme (Vlip509) showed the best hydrolyzing activity toward p-nitrophenyl butyrate (C(4)) among various p-nitrophenyl esters (C(2) to C(18)), and optimal activity of Vlip509 occurred at 30 degrees C and pH 8.5, respectively. Kinetic parameters toward p-nitrophenyl butyrate were determined as K (m) (307 muM), k (cat) (5.72 s(-1)), and k (cat)/K (m) (18.61 s(-1) mM(-1)). Furthermore, Vlip509 preferentially hydrolyzed the S-enantiomer of racemic ofloxacin ester. Despite its sequence homology to dienelactone hydrolase, Vlip509 showed no dienelactone hydrolase activity. This study represents the identification of a novel lipolytic enzyme from marine environment.
AuthorsSang-Yi Park, Jun-Tae Kim, Sung Gyun Kang, Jung-Hee Woo, Jung-Hyun Lee, Hyoung-Tae Choi, Sang-Jin Kim
JournalApplied microbiology and biotechnology (Appl Microbiol Biotechnol) Vol. 77 Issue 1 Pg. 107-15 (Nov 2007) ISSN: 0175-7598 [Print] Germany
PMID17712554 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
Chemical References
  • Bacterial Proteins
  • DNA, Bacterial
  • Recombinant Proteins
  • Esterases
  • Carboxylic Ester Hydrolases
  • carboxymethylenebutenolidase
Topics
  • Amino Acid Sequence
  • Bacterial Proteins (chemistry, genetics, metabolism)
  • Carboxylic Ester Hydrolases (chemistry, genetics, metabolism)
  • Cloning, Molecular
  • DNA, Bacterial (chemistry, genetics)
  • Electrophoresis, Polyacrylamide Gel
  • Esterases (chemistry, classification, metabolism)
  • Hydrogen-Ion Concentration
  • Molecular Sequence Data
  • Phylogeny
  • Recombinant Proteins (chemistry, isolation & purification, metabolism)
  • Sequence Analysis, DNA
  • Sequence Homology, Amino Acid
  • Stereoisomerism
  • Substrate Specificity
  • Temperature
  • Vibrio (classification, enzymology, genetics)

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