OHARA K, KUSADOKORO H, OZAKI J, HARUTA S, TAKAYA A, SAWAI T, MATSUMURA N, MINAMI S, IYOBE S; Interscience Conference on Antimicrobial Agents and Chemotherapy.

Abstr Intersci Conf Antimicrob Agents Chemother Intersci Conf Antimicrob Agents Chemother. 2000 Sep 17-20; 40: 101.

Chiba Univ., Chiba, Japan

beta-Lactamases are enzymes that hydrolyze beta-lactam antibiotics, conferring resistance to a variety of these antibiotics for most pathogenic bacteria. These enzymes have been classified phylogenetically based on their functional and molecular characteristics. Molecular class B metallo-beta-lactamase are characteristic in their broad substrate spectrum, which extend to most beta-lactam antibiotics, expect for monobactams, and have activities as penicillinases, cephalosporinases, and carbapenemases. In the course of surveying for the carbapenem-hydrolyzing metallo-beta-lactamase gene bla[IMP] in pathogenic bacteria by the PCR method, we detected a gene encoding a variant metallo-beta-lactamase, christened IMP-3, which differed from IMP-1 by having low hydrolyzing activity for penicillins and carbapenems. PCR direct-sequencing of a 2.2-kb segment revealed that the gene bla[IMP-3] was located on a cassette inserted within a class I integron in the pMS390 plasmid. The 741-bp nucleotide sequence of bla[IMP-3] was identical to that of bla[IMP-1], expect for seven base substitutions. Among these were, two, at nucleotide positions 314 and 640, which caused amino acid alterations. Hybrid bla genes were constructed from bla[IMP-3] and bla[IMP-1] by recombinant DNA technique, and beta-lactamases encoded by those genes were compared with those of the parents IMP-3 and IMP-1, under the same experimental conditions. The kinetic parameters indicated that the inefficient hydrolysis of benzylpenicillin, ampicillin, imipenem, and ceftazidime by IMP-3 was due to the substitution of glycine for serine at amino acid residue 196 in the mature enzyme. This alteration corresponded to the presence of guanine instead of an adenine at nucleotide position 640 of the bla[IMP-3] gene. This indicated that extension of the substrate profile in the metallo-beta-lactamase IMP-1 compared to IMP-3 is the result of a one-step single-base mutation, suggesting that the gene bla[IMP-3] is an ancestor of bla[IMP-1].KEYWORDS: Amino acid substitution; IMP-1; Metallo-beta-lactamase

Publication Types:

• Meeting Abstracts

Keywords:

• Amino Acid Substitution
• Anti-Bacterial Agents
• Base Sequence
• Carbapenems
• Imipenem
• Integrons
• Monobactams
• Plasmids
• Polymerase Chain Reaction
• beta-Lactamases
• beta-lactamase IMP-1
• genetics

Other ID:

• GWAIDS0011609

UI: 102249107

From Meeting Abstracts