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Crystal structures of KPC-2 ?-lactamase in complex with 3-nitrophenyl boronic acid and the penam sulfone PSR-3-226.

ABSTRACT: Class A carbapenemases are a major threat to the potency of carbapenem antibiotics. A widespread carbapenemase, KPC-2, is not easily inhibited by ?-lactamase inhibitors (i.e., clavulanic acid, sulbactam, and tazobactam). To explore different mechanisms of inhibition of KPC-2, we determined the crystal structures of KPC-2 with two ?-lactamase inhibitors that follow different inactivation pathways and kinetics. The first complex is that of a small boronic acid compound, 3-nitrophenyl boronic acid (3-NPBA), bound to KPC-2 with 1.62-Å resolution. 3-NPBA demonstrated a K(m) value of 1.0 ± 0.1 ?M (mean ± standard error) for KPC-2 and blocks the active site by making a reversible covalent interaction with the catalytic S70 residue. The two boron hydroxyl atoms of 3-NPBA are positioned in the oxyanion hole and the deacylation water pocket, respectively. In addition, the aromatic ring of 3-NPBA provides an edge-to-face interaction with W105 in the active site. The structure of KPC-2 with the penam sulfone PSR-3-226 was determined at 1.26-Å resolution. PSR-3-226 displayed a K(m) value of 3.8 ± 0.4 ?M for KPC-2, and the inactivation rate constant (k(inact)) was 0.034 ± 0.003 s(-1). When covalently bound to S70, PSR-3-226 forms a trans-enamine intermediate in the KPC-2 active site. The predominant active site interactions are generated via the carbonyl oxygen, which resides in the oxyanion hole, and the carboxyl moiety of PSR-3-226, which interacts with N132, N170, and E166. 3-NPBA and PSR-3-226 are the first ?-lactamase inhibitors to be trapped as an acyl-enzyme complex with KPC-2. The structural and inhibitory insights gained here could aid in the design of potent KPC-2 inhibitors.

SUBMITTER: Ke W

PROVIDER: S-EPMC3346646 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature

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Crystal structures of KPC-2 β-lactamase in complex with 3-nitrophenyl boronic acid and the penam sulfone PSR-3-226.

Ke Wei W Bethel Christopher R CR Papp-Wallace Krisztina M KM Pagadala Sundar Ram Reddy SR Nottingham Micheal M Fernandez Daniel D Buynak John D JD Bonomo Robert A RA van den Akker Focco F

Antimicrobial agents and chemotherapy 20120213 5

Class A carbapenemases are a major threat to the potency of carbapenem antibiotics. A widespread carbapenemase, KPC-2, is not easily inhibited by β-lactamase inhibitors (i.e., clavulanic acid, sulbactam, and tazobactam). To explore different mechanisms of inhibition of KPC-2, we determined the crystal structures of KPC-2 with two β-lactamase inhibitors that follow different inactivation pathways and kinetics. The first complex is that of a small boronic acid compound, 3-nitrophenyl boronic acid ...[more]

PMID: 22330909

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Project description:Resistance to expanded-spectrum cephalosporins and carbapenems has rendered certain strains of Klebsiella pneumoniae the most problematic pathogens infecting patients in the hospital and community. This broad-spectrum resistance to ?-lactamases emerges in part via the expression of KPC-2 and SHV-1 ?-lactamases and variants thereof. KPC-2 carbapenemase is particularly worrisome, as the genetic determinant encoding this ?-lactamase is rapidly spread via plasmids. Moreover, KPC-2, a class A enzyme, is difficult to inhibit with mechanism-based inactivators (e.g., clavulanate). In order to develop new ?-lactamase inhibitors (BLIs) to add to the limited available armamentarium that can inhibit KPC-2, we have structurally probed the boronic acid transition state analog S02030 for its inhibition of KPC-2 and SHV-1. S02030 contains a boronic acid, a thiophene, and a carboxyl triazole moiety. We present here the 1.54- and 1.87-Å resolution crystal structures of S02030 bound to SHV-1 and KPC-2 ?-lactamases, respectively, as well as a comparative analysis of the S02030 binding modes, including a previously determined S02030 class C ADC-7 ?-lactamase complex. S02030 is able to inhibit vastly different serine ?-lactamases by interacting with the conserved features of these active sites, which includes (i) forming the bond with catalytic serine via the boron atom, (ii) positioning one of the boronic acid oxygens in the oxyanion hole, and (iii) utilizing its amide moiety to make conserved interactions across the width of the active site. In addition, S02030 is able to overcome more distantly located structural differences between the ?-lactamases. This unique feature is achieved by repositioning the more polar carboxyl-triazole moiety, generated by click chemistry, to create polar interactions as well as reorient the more hydrophobic thiophene moiety. The former is aided by the unusual polar nature of the triazole ring, allowing it to potentially form a unique C-H…O 2.9-Å hydrogen bond with S130 in KPC-2.

Dataset Information

Crystal structures of KPC-2 ?-lactamase in complex with 3-nitrophenyl boronic acid and the penam sulfone PSR-3-226.

Publications

Crystal structures of KPC-2 β-lactamase in complex with 3-nitrophenyl boronic acid and the penam sulfone PSR-3-226.

Similar Datasets

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