Structural basis for the extended substrate spectrum of CMY‐10, a plasmid‐encoded class C β‐lactamase

• Published in: Molecular microbiology 2006-05, Vol.60 (4), p.907-916
• Main Authors: Kim, Jae Young, Jung, Ha Il, An, Young Jun, Lee, Jung Hun, Kim, So Jung, Jeong, Seok Hoon, Lee, Kye Joon, Suh, Pann‐Ghill, Lee, Heung‐Soo, Lee, Sang Hee, Cha, Sun‐Shin
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Anti-Bacterial Agents - chemistry; Anti-Bacterial Agents - metabolism; beta-Lactamases - chemistry; beta-Lactamases - genetics; Biological and medical sciences; Crystallography, X-Ray; Enterobacter cloacae; Enterobacter cloacae - enzymology; Enterobacter cloacae - genetics; Fundamental and applied biological sciences. Psychology; Hydrolysis; Imipenem - chemistry; Imipenem - metabolism; Microbiology; Molecular Sequence Data; Mutation; Plasmids - genetics; Protein Conformation; Sequence Deletion; Substrate Specificity
• ISSN: 0950-382X; 1365-2958
• DOI: 10.1111/j.1365-2958.2006.05146.x

Summary The emergence and dissemination of extended‐spectrum (ES) β‐lactamases induce therapeutic failure and a lack of eradication of clinical isolates even by third‐generation β‐lactam antibiotics like ceftazidime. CMY‐10 is a plasmid‐encoded class C β‐lactamase with a wide spectrum of substrates. Unlike the well‐studied class C ES β‐lactamase from Enterobacter cloacae GC1, the Ω‐loop does not affect the active site conformation and the catalytic activity of CMY‐10. Instead, a three‐amino‐acid deletion in the R2‐loop appears to be responsible for the ES activity of CMY‐10. According to the crystal structure solved at 1.55 Å resolution, the deletion significantly widens the R2 active site, which accommodates the R2 side‐chains of β‐lactam antibiotics. This observation led us to demonstrate the hydrolysing activity of CMY‐10 towards imipenem with a long R2 substituent. The forced mutational analyses of P99 β‐lactamase reveal that the introduction of deletion mutations into the R2‐loop is able to extend the substrate spectrum of class C non‐ES β‐lactamases, which is compatible with the isolation of natural class C ES enzymes harbouring deletion mutations in the R2‐loop. Consequently, the opening of the R2 active site by the deletion of some residues in the R2‐loop can be considered as an operative molecular strategy of class C β‐lactamases to extend their substrate spectrum.