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    Clinical variants of the native class d beta-lactamase of acinetobacter baumannii pose an emerging threat through ıncreased hydrolytic activity against carbapenems
    (Amer Soc Microbiology, 2016) Schroder, Emma C.; Klamer, Zachary L.; Saral, Ayşegül; Sugg, Kyle A.; June, Cynthia M.; Wymore, Troy; Szarecka, Agnieszka; Leonard, David A.
    The threat posed by the chromosomally encoded class D beta-lactamase of Acinetobacter baumannii (OXA-51/66) has been unclear, in part because of its relatively low affinity and turnover rate for carbapenems. Several hundred clinical variants of OXA51/66 have been reported, many with substitutions of active-site residues. We determined the kinetic properties of OXA-66 and five clinical variants with respect to a wide variety of beta-lactam substrates. The five variants displayed enhanced activity against carbapenems and in some cases against penicillins, late-generation cephalosporins, and the monobactam aztreonam. Molecular dynamics simulations show that in OXA-66, P130 inhibits the side-chain rotation of I129 and thereby prevents doripenem binding because of steric clash. A single amino acid substitution at this position (P130Q) in the variant OXA-109 greatly enhances the mobility of both I129 and a key active-site tryptophan (W222), thereby facilitating carbapenem binding. This expansion of substrate specificity represents a very worrisome development for the efficacy of beta-lactams against this troublesome pathogen.
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    Kinetic characterization of GES-22 beta-lactamase harboring the M169L clinical mutation
    (Japan Antibiotics Research Assoc, 2016) Saral, Ayşegül; Leonard, David A.; Düzgün, Azer Özad; Çopur Çiçek, Aysegül; June, Cynthia M.; Sandallı, Cemal
    The class A p-lactamase GES-22 has been identified in Acinetobacter baumannii isolates in Turkey, and subsequently shown to differ from GES-11 by a single substitution (M169L). Because M169 is part of the omega loop, a structure that is known to have major effects on substrate selectivity in class A beta-lactamases, we expressed, purified and kinetically characterized this novel variant. Our results show that compared with GES-11(6xHis), GES-22(6xHis) displays more efficient hydrolysis of penicillins, and aztreonam, but a loss of efficiency against ceftazidime. In addition, the M169L substitution confers on GES-22 more efficient hydrolysis of the mechanistic inhibitors clavulanic acid and sulbactam. These effects are highly similar to other mutations at the homologous position in other class A beta-lactamases, suggesting that this methionine has a key structural role in aligning active site residues and in substrate selectivity across the class.