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Cloning and expression of NDM-1 metallo-β-lactamase gene and study of the catalytic properties of the recombinant enzyme

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Abstract

A gene-expression system has been designed to express the NDM-1 metallo-β-lactamase gene in E. coli cells. This system enables the synthesis of the recombinant protein in a soluble and active form. A method for the isolation and purification of the recombinant enzyme has been developed. The yield of the homogeneous protein preparation was 10–15 mg per liter of E. coli culture medium. The catalytic parameters of the recombinant NDM-1 β-lactamase were measured for ampicillin (K m = 185 μM and k cat = 585 s–1) and meropenem (K m = 85 μM and k cat = 160 s–1). These values correlate well with the literature data. The catalytic parameters for the chromogenic CENTA substrate (K m = 14 μM and k cat = 290 s–1) were obtained for the first time.

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Authors and Affiliations

  1. Department of Chemistry, Moscow State University, 119991, Moscow, Russia

    V. G. Grigorenko, M. Yu. Rubtsova, E. V. Filatova, I. P. Andreeva, E. A. Mistryukova & A. M. Egorov

Authors
  1. V. G. Grigorenko
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  2. M. Yu. Rubtsova
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  3. E. V. Filatova
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  4. I. P. Andreeva
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  5. E. A. Mistryukova
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  6. A. M. Egorov
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Corresponding author

Correspondence to V. G. Grigorenko.

Additional information

Original Russian Text © V.G. Grigorenko, M.Yu. Rubtsova, E.V. Filatova, I.P. Andreeva, E.A. Mistryukova, A.M. Egorov, 2016, published in Vestnik Moskovskogo Universiteta. Khimiya, 2016, No. 2, pp. 75–81.

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Grigorenko, V.G., Rubtsova, M.Y., Filatova, E.V. et al. Cloning and expression of NDM-1 metallo-β-lactamase gene and study of the catalytic properties of the recombinant enzyme. Moscow Univ. Chem. Bull. 71, 104–109 (2016). https://doi.org/10.3103/S0027131416020048

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  • DOI: https://doi.org/10.3103/S0027131416020048

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