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Cloning and sequence analysis of the gene for a carbapenem-hydrolyzing class A beta-lactamase, Sme-1, from Serratia marcescens S6
Author(s) -
Thierry Naas,
Laurence Vandel,
Wladimir Sougakoff,
David M. Livermore,
Patrice Nordmann
Publication year - 1994
Publication title -
antimicrobial agents and chemotherapy
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.07
H-Index - 259
eISSN - 1070-6283
pISSN - 0066-4804
DOI - 10.1128/aac.38.6.1262
Subject(s) - serratia marcescens , biology , enterobacter cloacae , peptide sequence , open reading frame , escherichia coli , carbapenem , plasmid , microbiology and biotechnology , beta lactamase , gene , enterobacteriaceae , genetics , antibiotics
Serratia marcescens S6 produces a pI 9.7 carbapenem-hydrolyzing beta-lactamase that is probably encoded by the chromosome (Y. Yang, P. Wu, and D. M. Livermore, Antimicrob. Agents Chemother. 34:755-758, 1990). A total of 11.3 kb of genomic DNA from this strain was cloned into plasmid pACYC184 in Escherichia coli. After further subclonings, the carbapenem-hydrolyzing beta-lactamase gene (blaSme-1) was sequenced (EMBL accession number Z28968). The gene corresponded to an 882-bp open reading frame which encoded a 294-amino-acid polypeptide. This open reading frame was preceded by a -10 and a -35 region consistent with a putative promoter sequence of members of the family Enterobacteriaceae. This promoter was active in E. coli and S. marcescens, as demonstrated by primer extension analysis. N-terminal sequencing showed that the Sme-1 enzyme had a 27-amino-acid leader peptide and enabled calculation of the molecular mass of the mature protein (29.3 kDa). Sequence alignment revealed that Sme-1 is a class A serine beta-lactamase and not a class B metalloenzyme. The earlier view that the enzyme was zinc dependent was discounted. Among class A beta-lactamases, Sme-1 had the greatest amino acid identity (70%) with the pI 6.9 carbapenem-hydrolyzing beta-lactamase, NMC-A, from Enterobacter cloacae NOR-1. Comparison of these two protein sequences suggested a role for specific residues in carbapenem hydrolysis. The relatedness of Sme-1 to other class A beta-lactamases such as the TEM and SHV types was remote. This work details the sequence of the second carbapenem-hydrolyzing class A beta-lactamase from an enterobacterial species and the first in the genus Serratia.

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