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A new acylamidase from Rhodococcus erythropolis TA37 can hydrolyze N-substituted amides

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Abstract

A new acylamidase was isolated from Rhodococcus erythropolis TA37 and characterized. N-Substituted acrylamides (isopropyl acrylamide, N,N-dimethyl-aminopropyl acrylamide, and methylene-bis-acrylamide), acid para-nitroanilides (4′-nitroacetanilide, Gly-pNA, Ala-pNA, Leu-pNA), and N-acetyl derivatives of glycine, alanine, and leucine are good substrates for this enzyme. Aliphatic amides (acetamide, acrylamide, isobutyramide, n-butyramide, and valeramide) are also used as substrates but with less efficiency. The enzyme subunit mass by SDS-PAGE is 55 kDa. Maximal activity is exhibited at pH 7–8 and 55°C. The enzyme is stable for 15 h at 22°C and for 0.5 h at 45°C. The Michaelis constant (K m) is 0.25 mM with Gly-pNA and 0.55 mM with Ala-pNA. The acylamidase activity is suppressed by inhibitors of serine proteases (phenylmethylsulfonyl fluoride and diisopropyl fluorophosphate) but is not suppressed by inhibitors of aliphatic amidases (acetaldehyde and nitrophenyl disulfides). The N-terminal amino acid sequence of the acylamidase is highly homologous to those of two putative amidases detected from sequenced R. erythropolis genomes. It is suggested that the acylamidase together with the detected homologs forms a new class within the amidase signature family.

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Abbreviations

Ala-pNA:

alanine para-nitroanilide

DIFP:

diisopropyl fluorophosphate

Gly-pNA:

glycine para-nitroanilide

Leu-pNA:

leucine para-nitroanilide

NAA:

4′-nitroacetanilide

PMSF:

phenylmethylsulfonyl fluoride

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Authors and Affiliations

  1. Institute for Genetics and Selection of Industrial Microorganisms, 1-yi Dorozhnyi Proezd 1, 113545, Moscow, Russia

    K. V. Lavrov, I. A. Zalunin, E. K. Kotlova & A. S. Yanenko

Authors
  1. K. V. Lavrov
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  2. I. A. Zalunin
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  3. E. K. Kotlova
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  4. A. S. Yanenko
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Corresponding author

Correspondence to A. S. Yanenko.

Additional information

Original Russian Text © K. V. Lavrov, I. A. Zalunin, E. K. Kotlova, A. S. Yanenko, 2010, published in Biokhimiya, 2010, Vol. 75, No. 8, pp. 1111–1119.

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Lavrov, K.V., Zalunin, I.A., Kotlova, E.K. et al. A new acylamidase from Rhodococcus erythropolis TA37 can hydrolyze N-substituted amides. Biochemistry Moscow 75, 1006–1013 (2010). https://doi.org/10.1134/S0006297910080080

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  • DOI: https://doi.org/10.1134/S0006297910080080

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