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V. G. Grigorenko, M. Yu. Rubtsova, E. V. Filatova, I. P. Andreeva, E. A. Mistryukova, A. M. Egorov
Cloning, expression of
NDM-1 metallo-β-lactamase gene and study of catalytic properties of the
recombinant enzyme
Abstract
The
gene expression system of NDM-1 metallo-β-lactamase in E. coli cells,
which provides a synthesis of a recombinant protein in soluble, active form,
has been developed. A method for the isolation and purification of the
recombinant enzyme allowed for homogeneous protein preparation to 10–15 mg from
1 liter of E. coli culture medium. Kinetic parameters for recombinant
NDM-1 β-lactamase measured for ampicillin (KM eff= 185 µM, kcat= 585 s–1) and
meropenem (KM eff = 85 µM, kcat= 160 s–1)
correlate well with literature data. For the first kinetic parameters have been
obtained for chromogenic substrate CENTA: KM eff = 14 µM, kcat= 290 s–1.
Key words: recombinant metallo-β-lactamase
NDM-1, ampicillin, meropenem, CENTA, kinetic parameters.
Copyright (C) Chemistry Dept., Moscow State University, 2002
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