Cloning and sequence analysis of the gene for a carbapenem-hydrolyzing class A beta-lactamase, Sme-1, from Serratia marcescens S6.
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.
Naas
T
T
Abteilung Mikrobiologie, Universität Basel, Switzerland.
Vandel
L
L
Sougakoff
W
W
Livermore
D M
DM
Nordmann
P
P
D000595:Amino Acid Sequence; D001483:Base Sequence; D015780:Carbapenems / Q000378:metabolism*; D003001:Cloning, Molecular; D005798:Genes, Bacterial; D006868:Hydrolysis; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D012706:Serratia marcescens / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D001618:beta-Lactamases / Q000737:chemistry / Q000235:genetics*
D015780:Carbapenems; C094219:beta-lactamase Sme-1; D001618:beta-Lactamases