Conformational flexibility of the glycosidase NagZ allows it to bind structurally diverse inhibitors to suppress β‐lactam antibiotic resistance
NagZ is an N‐acetyl‐β‐d‐glucosaminidase that participates in the peptidoglycan (PG) recycling pathway of Gram‐negative bacteria by removing N‐acetyl‐glucosamine (GlcNAc) from PG fragments that have been excised from the cell wall during growth. The 1,6‐anhydromuramoyl‐peptide products generated by N...
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Published in | Protein science Vol. 26; no. 6; pp. 1161 - 1170 |
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Main Authors | , , , , , |
Format | Journal Article |
Language | English |
Published |
United States
Wiley Subscription Services, Inc
01.06.2017
Wiley John Wiley and Sons Inc |
Subjects | |
Online Access | Get full text |
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Summary: | NagZ is an N‐acetyl‐β‐d‐glucosaminidase that participates in the peptidoglycan (PG) recycling pathway of Gram‐negative bacteria by removing N‐acetyl‐glucosamine (GlcNAc) from PG fragments that have been excised from the cell wall during growth. The 1,6‐anhydromuramoyl‐peptide products generated by NagZ activate β‐lactam resistance in many Gram‐negative bacteria by inducing the expression of AmpC β‐lactamase. Blocking NagZ activity can thereby suppress β‐lactam antibiotic resistance in these bacteria. The NagZ active site is dynamic and it accommodates distortion of the glycan substrate during catalysis using a mobile catalytic loop that carries a histidine residue which serves as the active site general acid/base catalyst. Here, we show that flexibility of this catalytic loop also accommodates structural differences in small molecule inhibitors of NagZ, which could be exploited to improve inhibitor specificity. X‐ray structures of NagZ bound to the potent yet non‐selective N‐acetyl‐β‐glucosaminidase inhibitor PUGNAc (O‐(2‐acetamido‐2‐deoxy‐d‐glucopyranosylidene) amino‐N‐phenylcarbamate), and two NagZ‐selective inhibitors – EtBuPUG, a PUGNAc derivative bearing a 2‐N‐ethylbutyryl group, and MM‐156, a 3‐N‐butyryl trihydroxyazepane, revealed that the phenylcarbamate moiety of PUGNAc and EtBuPUG completely displaces the catalytic loop from the NagZ active site to yield a catalytically incompetent form of the enzyme. In contrast, the catalytic loop was found positioned in the catalytically active conformation within the NagZ active site when bound to MM‐156, which lacks the phenylcarbamate extension. Displacement of the catalytic loop by PUGNAc and its N‐acyl derivative EtBuPUG alters the active site conformation of NagZ, which presents an additional strategy to improve the potency and specificity of NagZ inhibitors.
PDB Code(s): 5UTQ; 5UTP; 5UTR |
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Bibliography: | NagZ is an enzyme within the peptidoglycan recycling pathway required for the activation of AmpC β‐lactamase production in several pathogenic Gram‐negative bacteria. AmpC confers resistance to numerous clinically important β‐lactam antibiotics. Thus, blocking NagZ activity with small‐molecule inhibitors has garnered interest as a therapeutic strategy to suppress AmpC‐mediated β‐lactam resistance. Here, we provide a clear structural framework for improving the potency and selectivity of NagZ inhibitors for use in enhancing the efficacy of β‐lactam antibiotics. Statement ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 PMCID: PMC5441427 Statement: NagZ is an enzyme within the peptidoglycan recycling pathway required for the activation of AmpC β‐lactamase production in several pathogenic Gram‐negative bacteria. AmpC confers resistance to numerous clinically important β‐lactam antibiotics. Thus, blocking NagZ activity with small‐molecule inhibitors has garnered interest as a therapeutic strategy to suppress AmpC‐mediated β‐lactam resistance. Here, we provide a clear structural framework for improving the potency and selectivity of NagZ inhibitors for use in enhancing the efficacy of β‐lactam antibiotics. |
ISSN: | 0961-8368 1469-896X |
DOI: | 10.1002/pro.3166 |