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A membrane-bound esterase PA2949 from Pseudomonas aeruginosa is expressed and purified from Escherichia coli.

ABSTRACT: Pseudomonas aeruginosa strain 1001 produces an esterase (EstA) that can hydrolyse the racemic methyl ester of ?-acetylthioisobutyrate to produce the (D)-enantiomer, which serves as a precursor of captopril, a drug used for treatment of hypertension. We show here that PA2949 from P. aeruginosa PA01, a homologue of EstA, can efficiently be expressed in an enzymatically active form in E. coli. The enzyme is membrane-associated as demonstrated by cell fractionation studies. PA2949 was purified to homogeneity after solubilisation with the nonionic detergent, Triton X-100, and was shown to possess a conserved esterase catalytic triad consisting of Ser137-His258-Asp286. Our results should allow the development of an expression and purification strategy to produce this biotechnologically relevant esterase in a pure form with a high yield.

SUBMITTER: Kovacic F

PROVIDER: S-EPMC4856427 | biostudies-literature | 2016 May

REPOSITORIES: biostudies-literature

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A membrane-bound esterase PA2949 from Pseudomonas aeruginosa is expressed and purified from Escherichia coli.

Kovacic Filip F Bleffert Florian F Caliskan Muttalip M Wilhelm Susanne S Granzin Joachim J Batra-Safferling Renu R Jaeger Karl-Erich KE

FEBS open bio 20160419 5

Pseudomonas aeruginosa strain 1001 produces an esterase (EstA) that can hydrolyse the racemic methyl ester of β-acetylthioisobutyrate to produce the (D)-enantiomer, which serves as a precursor of captopril, a drug used for treatment of hypertension. We show here that PA2949 from P. aeruginosa PA01, a homologue of EstA, can efficiently be expressed in an enzymatically active form in E. coli. The enzyme is membrane-associated as demonstrated by cell fractionation studies. PA2949 was purified to ho ...[more]

PMID: 27419054

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