Andreas Prester, Markus Perbandt, Marina Galchenkova, Dominik Oberthuer, Nadine Werner, Alessandra Henkel, Julia Maracke, Oleksandr Yefanov, Johanna Hakanpää, Guillaume Pompidor, Jan Meyer, Henry Chapman, Martin Aepfelbacher, Winfried Hinrichs, Holger Rohde, Christian Betzel
{"title":"Time-resolved crystallography of boric acid binding to the active site serine of the β-lactamase CTX-M-14 and subsequent 1,2-diol esterification","authors":"Andreas Prester, Markus Perbandt, Marina Galchenkova, Dominik Oberthuer, Nadine Werner, Alessandra Henkel, Julia Maracke, Oleksandr Yefanov, Johanna Hakanpää, Guillaume Pompidor, Jan Meyer, Henry Chapman, Martin Aepfelbacher, Winfried Hinrichs, Holger Rohde, Christian Betzel","doi":"10.1038/s42004-024-01236-w","DOIUrl":null,"url":null,"abstract":"The emergence and spread of antibiotic resistance represent a growing threat to public health. Of particular concern is the appearance of β-lactamases, which are capable to hydrolyze and inactivate the most important class of antibiotics, the β-lactams. Effective β-lactamase inhibitors and mechanistic insights into their action are central in overcoming this type of resistance, and in this context boronate-based β-lactamase inhibitors were just recently approved to treat multidrug-resistant bacteria. Using boric acid as a simplified inhibitor model, time-resolved serial crystallography was employed to obtain mechanistic insights into binding to the active site serine of β-lactamase CTX-M-14, identifying a reaction time frame of 80–100 ms. In a next step, the subsequent 1,2-diol boric ester formation with glycerol in the active site was monitored proceeding in a time frame of 100–150 ms. Furthermore, the displacement of the crucial anion in the active site of the β-lactamase was verified as an essential part of the binding mechanism of substrates and inhibitors. In total, 22 datasets of β-lactamase intermediate complexes with high spatial resolution of 1.40–2.04 Å and high temporal resolution range of 50–10,000 ms were obtained, allowing a detailed analysis of the studied processes. Mechanistic details captured here contribute to the understanding of molecular processes and their time frames in enzymatic reactions. Moreover, we could demonstrate that time-resolved crystallography can serve as an additional tool for identifying and investigating enzymatic reactions. Boronate-based ß-lactamase inhibitors play an important role in treating multidrug-resistant bacteria infection, however, the molecular mechanism of inhibition remains unclear. Here, the authors use time-resolved serial crystallography to investigate the binding process by using boric acid as a model against β-lactamase CTX-M-14, revealing the binding to the active site serine within 80–100 ms, a subsequent 1,2-diol boric ester formation with glycerol within 100–150 ms, as well as the displacement of the sulfate anion in the active site.","PeriodicalId":10529,"journal":{"name":"Communications Chemistry","volume":null,"pages":null},"PeriodicalIF":5.9000,"publicationDate":"2024-07-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.nature.com/articles/s42004-024-01236-w.pdf","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Communications Chemistry","FirstCategoryId":"92","ListUrlMain":"https://www.nature.com/articles/s42004-024-01236-w","RegionNum":2,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"CHEMISTRY, MULTIDISCIPLINARY","Score":null,"Total":0}
引用次数: 0
Abstract
The emergence and spread of antibiotic resistance represent a growing threat to public health. Of particular concern is the appearance of β-lactamases, which are capable to hydrolyze and inactivate the most important class of antibiotics, the β-lactams. Effective β-lactamase inhibitors and mechanistic insights into their action are central in overcoming this type of resistance, and in this context boronate-based β-lactamase inhibitors were just recently approved to treat multidrug-resistant bacteria. Using boric acid as a simplified inhibitor model, time-resolved serial crystallography was employed to obtain mechanistic insights into binding to the active site serine of β-lactamase CTX-M-14, identifying a reaction time frame of 80–100 ms. In a next step, the subsequent 1,2-diol boric ester formation with glycerol in the active site was monitored proceeding in a time frame of 100–150 ms. Furthermore, the displacement of the crucial anion in the active site of the β-lactamase was verified as an essential part of the binding mechanism of substrates and inhibitors. In total, 22 datasets of β-lactamase intermediate complexes with high spatial resolution of 1.40–2.04 Å and high temporal resolution range of 50–10,000 ms were obtained, allowing a detailed analysis of the studied processes. Mechanistic details captured here contribute to the understanding of molecular processes and their time frames in enzymatic reactions. Moreover, we could demonstrate that time-resolved crystallography can serve as an additional tool for identifying and investigating enzymatic reactions. Boronate-based ß-lactamase inhibitors play an important role in treating multidrug-resistant bacteria infection, however, the molecular mechanism of inhibition remains unclear. Here, the authors use time-resolved serial crystallography to investigate the binding process by using boric acid as a model against β-lactamase CTX-M-14, revealing the binding to the active site serine within 80–100 ms, a subsequent 1,2-diol boric ester formation with glycerol within 100–150 ms, as well as the displacement of the sulfate anion in the active site.
期刊介绍:
Communications Chemistry is an open access journal from Nature Research publishing high-quality research, reviews and commentary in all areas of the chemical sciences. Research papers published by the journal represent significant advances bringing new chemical insight to a specialized area of research. We also aim to provide a community forum for issues of importance to all chemists, regardless of sub-discipline.