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Structural and mechanistic insights into the inhibition of class C beta-lactamases through the adenylylation of the nucleophilic serine

Title
Structural and mechanistic insights into the inhibition of class C beta-lactamases through the adenylylation of the nucleophilic serine
Authors
Kim, Min-KyuAn, Young JunNa, Jung-HyunSeol, Jae-HeeRyu, Ju YeonLee, Jin-WonKang, Lin-WooChung, Kyung MinLee, Jung-HyunMoon, Jeong HeeLee, Jong SeokCha, Sun-Shin
Ewha Authors
차선신
SCOPUS Author ID
차선신scopus
Issue Date
2017
Journal Title
JOURNAL OF ANTIMICROBIAL CHEMOTHERAPY
ISSN
0305-7453JCR Link1460-2091JCR Link
Citation
vol. 72, no. 3, pp. 735 - 743
Publisher
OXFORD UNIV PRESS
Indexed
SCI; SCIE; SCOPUS WOS
Abstract
Objectives: Investigation into the adenylylation of the nucleophilic serine in AmpC BER and CMY-10 extended spectrumclass C beta-lactamases. Methods: The formation and the stability of the adenylate adduct were examined by X-ray crystallography and MS. Inhibition assays for kinetic parameters were performed by monitoring the hydrolytic activity of AmpC BER and CMY-10 using nitrocefin as a reporter substrate. The effect of adenosine 50'-(P-acetyl) monophosphate ( acAMP) on the MIC of ceftazidime was tested with four Gram-negative clinical isolates. Results: The crystal structures and MS analyses confirmed the acAMP-mediated adenylylation of the nucleophilic serine in AmpC BER and CMY-10. acAMP inhibited AmpC BER and CMY-10 through the adenylylation of the nucleophilic serine, which could bemodelled as a two-stepmechanism. The initial non-covalent binding of acAMP to the active site is followed by the covalent attachment of its AMP moiety to the nucleophilic serine. The inhibition efficiencies (k(inact)/K-I) of acAMP against AmpC BER and CMY-10 were determined to be 320 and 140 M(-1)s(-1), respectively. The combination of ceftazidime and acAMP reduced the MIC of ceftazidime against the tested bacteria. Conclusions: Our structural and kinetic studies revealed the detailed mechanismof adenylylation of the nucleophilic serine and may serve as a starting point for the design of novel class C beta-lactamase inhibitors on the basis of the nucleotide scaffold.
DOI
10.1093/jac/dkw491
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자연과학대학 > 화학·나노과학전공 > Journal papers
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