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PLoS One. 2014 Jan 16;9(1):e85892. doi: 10.1371/journal.pone.0085892. eCollection 2014.

Penam sulfones and β-lactamase inhibition: SA2-13 and the importance of the C2 side chain length and composition.

Author information

1
Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio, United States of America.
2
Department of Molecular Biology and Microbiology, Case Western Reserve University, Cleveland, Ohio, United States of America ; Research Division, Louis Stokes Cleveland Veterans Affairs Medical Center, Cleveland, Ohio, United States of America.
3
Research Division, Louis Stokes Cleveland Veterans Affairs Medical Center, Cleveland, Ohio, United States of America.
4
Department of Chemistry, Southern Methodist University, Dallas, Texas, United States of America.
5
Department of Molecular Biology and Microbiology, Case Western Reserve University, Cleveland, Ohio, United States of America ; Research Division, Louis Stokes Cleveland Veterans Affairs Medical Center, Cleveland, Ohio, United States of America ; Department of Medicine, Case Western Reserve University, Cleveland, Ohio, United States of America ; Department of Pharmacology, Case Western Reserve University, Cleveland, Ohio, United States of America.

Abstract

β-Lactamases are the major reason β-lactam resistance is seen in Gram-negative bacteria. To combat this resistance mechanism, β-lactamase inhibitors are currently being developed. Presently, there are only three that are in clinical use (clavulanate, sulbactam and tazobactam). In order to address this important medical need, we explored a new inhibition strategy that takes advantage of a long-lived inhibitory trans-enamine intermediate. SA2-13 was previously synthesized and shown to have a lower k(react) than tazobactam. We investigated here the importance of the carboxyl linker length and composition by synthesizing three analogs of SA2-13 (PSR-4-157, PSR-4-155, and PSR-3-226). All SA2-13 analogs yielded higher turnover numbers and k(react) compared to SA2-13. We next demonstrated using protein crystallography that increasing the linker length by one carbon allowed for better capture of a trans-enamine intermediate; in contrast, this trans-enamine intermediate did not occur when the C2 linker length was decreased by one carbon. If the linker was altered by both shortening it and changing the carboxyl moiety into a neutral amide moiety, the stable trans-enamine intermediate in wt SHV-1 did not form; this intermediate could only be observed when a deacylation deficient E166A variant was studied. We subsequently studied SA2-13 against a relatively recently discovered inhibitor-resistant (IR) variant of SHV-1, SHV K234R. Despite the alteration in the mechanism of resistance due to the K→R change in this variant, SA2-13 was effective at inhibiting this IR enzyme and formed a trans-enamine inhibitory intermediate similar to the intermediate seen in the wt SHV-1 structure. Taken together, our data reveals that the C2 side chain linker length and composition profoundly affect the formation of the trans-enamine intermediate of penam sulfones. We also show that the design of SA2-13 derivatives offers promise against IR SHV β-lactamases that possess the K234R substitution.

PMID:
24454944
PMCID:
PMC3894197
DOI:
10.1371/journal.pone.0085892
[Indexed for MEDLINE]
Free PMC Article

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