Insights into Class D β-Lactamases Are Revealed by the Crystal Structure of the OXA10 Enzyme from Pseudomonas aeruginosa

• Published in: Structure (London) 2000-12, Vol.8 (12), p.1289-1298
• Main Authors: Maveyraud, Laurent, Golemi, Dasantila, Kotra, Lakshmi P., Tranier, Samuel, Vakulenko, Sergei, Mobashery, Shahriar, Samama, Jean-Pierre
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; antibiotic resistance; Bacterial Proteins; beta-Lactam Resistance; beta-Lactamases - chemistry; beta-Lactamases - metabolism; beta-Lactamases - pharmacology; Binding Sites; Carrier Proteins - antagonists & inhibitors; Catalysis; crystal structure; Crystallization; Crystallography, X-Ray; Dimerization; Evolution, Molecular; Hexosyltransferases; MAD; Molecular Sequence Data; Muramoylpentapeptide Carboxypeptidase - antagonists & inhibitors; Penicillin-Binding Proteins; Penicillins - metabolism; Peptidyl Transferases; Protein Structure, Secondary; Protein Structure, Tertiary; Pseudomonas aeruginosa - drug effects; Pseudomonas aeruginosa - enzymology; Sequence Alignment; Sequence Homology, Amino Acid; β-lactamase
• ISSN: 0969-2126; 1878-4186
• DOI: 10.1016/S0969-2126(00)00534-7

Background: β-lactam antibiotic therapies are commonly challenged by the hydrolytic activities of β-lactamases in bacteria. These enzymes have been grouped into four classes: A, B, C, and D. Class B β-lactamases are zinc dependent, and enzymes of classes A, C, and D are transiently acylated on a serine residue in the course of the turnover chemistry. While class A and C β-lactamases have been extensively characterized by biochemical and structural methods, class D enzymes remain the least studied despite their increasing importance in the clinic. Results: The crystal structure of the OXA10 class D β-lactamase has been solved to 1.66 Å resolution from a gold derivative and MAD phasing. This structure reveals that β-lactamases from classes D and A, despite very poor sequence similarity, share a similar overall fold. An additional β strand in OXA10 mediates the association into dimers characterized by analytical ultracentrifugation. Major differences are found when comparing the molecular details of the active site of this class D enzyme to the corresponding regions in class A and C β-lactamases. In the native structure of the OXA10 enzyme solved to 1.8 Å, Lys-70 is carbamylated. Conclusions: Several features were revealed by this study: the dimeric structure of the OXA10 β-lactamase, an extension of the substrate binding site which suggests that class D enzymes may bind other substrates beside β-lactams, and carbamylation of the active site Lys-70 residue. The CO 2-dependent activity of the OXA10 enzyme and the kinetic properties of the natural OXA17 mutant protein suggest possible relationships between carbamylation, inhibition of the enzyme by anions, and biphasic behavior of the enzyme.