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Identifying antibiotics based on structural differences in the conserved allostery from mitochondrial heme-copper oxidases.

ABSTRACT: Antimicrobial resistance (AMR) is a global health problem. Despite the enormous efforts made in the last decade, threats from some species, including drug-resistant Neisseria gonorrhoeae, continue to rise and would become untreatable. The development of antibiotics with a different mechanism of action is seriously required. Here, we identified an allosteric inhibitory site buried inside eukaryotic mitochondrial heme-copper oxidases (HCOs), the essential respiratory enzymes for life. The steric conformation around the binding pocket of HCOs is highly conserved among bacteria and eukaryotes, yet the latter has an extra helix. This structural difference in the conserved allostery enabled us to rationally identify bacterial HCO-specific inhibitors: an antibiotic compound against ceftriaxone-resistant Neisseria gonorrhoeae. Molecular dynamics combined with resonance Raman spectroscopy and stopped-flow spectroscopy revealed an allosteric obstruction in the substrate accessing channel as a mechanism of inhibition. Our approach opens fresh avenues in modulating protein functions and broadens our options to overcome AMR.

SUBMITTER: Nishida Y

PROVIDER: S-EPMC9731990 | biostudies-literature | 2022 Dec

REPOSITORIES: biostudies-literature

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Identifying antibiotics based on structural differences in the conserved allostery from mitochondrial heme-copper oxidases.

Nishida Yuya Y Yanagisawa Sachiko S Morita Rikuri R Shigematsu Hideki H Shinzawa-Itoh Kyoko K Yuki Hitomi H Ogasawara Satoshi S Shimuta Ken K Iwamoto Takashi T Nakabayashi Chisa C Matsumura Waka W Kato Hisakazu H Gopalasingam Chai C Nagao Takemasa T Qaqorh Tasneem T Takahashi Yusuke Y Yamazaki Satoru S Kamiya Katsumasa K Harada Ryuhei R Mizuno Nobuhiro N Takahashi Hideyuki H Akeda Yukihiro Y Ohnishi Makoto M Ishii Yoshikazu Y Kumasaka Takashi T Murata Takeshi T Muramoto Kazumasa K Tosha Takehiko T Shiro Yoshitsugu Y Honma Teruki T Shigeta Yasuteru Y Kubo Minoru M Takashima Seiji S Shintani Yasunori Y

Nature communications 20221208 1

Antimicrobial resistance (AMR) is a global health problem. Despite the enormous efforts made in the last decade, threats from some species, including drug-resistant Neisseria gonorrhoeae, continue to rise and would become untreatable. The development of antibiotics with a different mechanism of action is seriously required. Here, we identified an allosteric inhibitory site buried inside eukaryotic mitochondrial heme-copper oxidases (HCOs), the essential respiratory enzymes for life. The steric c ...[more]

PMID: 36481732

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Project description:The route of O₂to and from the high-spin heme in heme-copper oxidases has generally been believed to emulate that of carbon monoxide (CO). Time-resolved and stationary infrared experiments in our laboratories of the fully reduced CO-bound enzymes, as well as transient optical absorption saturation kinetics studies as a function of CO pressure, have provided strong support for CO binding to CuB⁺ on the pathway to and from the high-spin heme. The presence of CO on CuB⁺ suggests that O₂binding may be compromised in CO flow-flash experiments. Time-resolved optical absorption studies show that the rate of O₂and NO binding in the bovine enzyme (1 × 10⁸M⁻¹s⁻¹) is unaffected by the presence of CO, which is consistent with the rapid dissociation (t½ = 1.5μs) of CO from CuB⁺. In contrast, in Thermus thermophilus (Tt) cytochrome ba3 the O₂and NO binding to heme a3 slows by an order of magnitude in the presence of CO (from 1 × 10⁹ to 1 × 10⁸M⁻¹s⁻¹), but is still considerably faster (~10μs at 1atm O₂) than the CO off-rate from CuB in the absence of O₂(milliseconds). These results show that traditional CO flow-flash experiments do not give accurate results for the physiological binding of O₂and NO in Tt ba3, namely, in the absence of CO. They also raise the question whether in CO flow-flash experiments on Tt ba3 the presence of CO on CuB⁺ impedes the binding of O₂to CuB⁺ or, if O₂does not bind to CuB⁺ prior to heme a3, whether the CuB⁺-CO complex sterically restricts access of O₂to the heme. Both possibilities are discussed, and we argue that O₂binds directly to heme a3 in Tt ba3, causing CO to dissociate from CuB⁺ in a concerted manner through steric and/or electronic effects. This would allow CuB⁺ to function as an electron donor during the fast (5μs) breaking of the OO bond. These results suggest that the binding of CO to CuB⁺ on the path to and from heme a3 may not be applicable to O₂and NO in all heme-copper oxidases. This article is part of a Special Issue entitled: Vibrational spectroscopies and bioenergetic systems.

Dataset Information

Identifying antibiotics based on structural differences in the conserved allostery from mitochondrial heme-copper oxidases.

Publications

Identifying antibiotics based on structural differences in the conserved allostery from mitochondrial heme-copper oxidases.

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