Structural and Functional Characterization of OXA-48: Insight into Mechanism and Structural Basis of Substrate Recognition and Specificity

Class D β-lactamase OXA-48 is widely distributed among Gram-negative bacteria and is an important determinant of resistance to the last-resort carbapenems. Nevertheless, the detailed mechanism by which this β-lactamase hydrolyzes its substrates remains poorly understood. In this study, the complex s...

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Autores principales:	Jiachi Chiou, Qipeng Cheng, Perry Tim-fat Shum, Marcus Ho-yin Wong, Edward Wai-chi Chan, Sheng Chen
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	carbapenemase OXA-48 active site residues interaction β-lactams Biology (General) QH301-705.5 Chemistry QD1-999
Acceso en línea:	https://doaj.org/article/f47b3272ce3a49a29acb1e096c33c58c
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spelling	oai:doaj.org-article:f47b3272ce3a49a29acb1e096c33c58c2021-11-11T16:56:59ZStructural and Functional Characterization of OXA-48: Insight into Mechanism and Structural Basis of Substrate Recognition and Specificity10.3390/ijms2221114801422-00671661-6596https://doaj.org/article/f47b3272ce3a49a29acb1e096c33c58c2021-10-01T00:00:00Zhttps://www.mdpi.com/1422-0067/22/21/11480https://doaj.org/toc/1661-6596https://doaj.org/toc/1422-0067Class D β-lactamase OXA-48 is widely distributed among Gram-negative bacteria and is an important determinant of resistance to the last-resort carbapenems. Nevertheless, the detailed mechanism by which this β-lactamase hydrolyzes its substrates remains poorly understood. In this study, the complex structures of OXA-48 and various β-lactams were modeled and the potential active site residues that may interact with various β-lactams were identified and characterized to elucidate their roles in OXA-48 substrate recognition. Four residues, namely S<sup>70</sup>, K<sup>73</sup>, S<sup>118</sup>, and K<sup>208</sup> were found to be essential for OXA-48 to undergo catalytic hydrolysis of various penicillins and carbapenems both in vivo and in vitro. T<sup>209</sup> was found to be important for hydrolysis of imipenem, whereas R<sup>250</sup> played a major role in hydrolyzing ampicillin, imipenem, and meropenem most likely by forming a H-bond or salt-bridge between the side chain of these two residues and the carboxylate oxygen ions of the substrates. Analysis of the effect of substitution of alanine in two residues, W<sup>105</sup> and L<sup>158</sup>, revealed their roles in mediating the activity of OXA-48. Our data show that these residues most likely undergo hydrophobic interaction with the R groups and the core structure of the β-lactam ring in penicillins and the carbapenems, respectively. Unlike OXA-58, mass spectrometry suggested a loss of the C6-hydroxyethyl group during hydrolysis of meropenem by OXA-48, which has never been demonstrated in Class D carbapenemases. Findings in this study provide comprehensive knowledge of the mechanism of the substrate recognition and catalysis of OXA-type β-lactamases.Jiachi ChiouQipeng ChengPerry Tim-fat ShumMarcus Ho-yin WongEdward Wai-chi ChanSheng ChenMDPI AGarticlecarbapenemaseOXA-48active site residuesinteractionβ-lactamsBiology (General)QH301-705.5ChemistryQD1-999ENInternational Journal of Molecular Sciences, Vol 22, Iss 11480, p 11480 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	carbapenemase OXA-48 active site residues interaction β-lactams Biology (General) QH301-705.5 Chemistry QD1-999
spellingShingle	carbapenemase OXA-48 active site residues interaction β-lactams Biology (General) QH301-705.5 Chemistry QD1-999 Jiachi Chiou Qipeng Cheng Perry Tim-fat Shum Marcus Ho-yin Wong Edward Wai-chi Chan Sheng Chen Structural and Functional Characterization of OXA-48: Insight into Mechanism and Structural Basis of Substrate Recognition and Specificity
description	Class D β-lactamase OXA-48 is widely distributed among Gram-negative bacteria and is an important determinant of resistance to the last-resort carbapenems. Nevertheless, the detailed mechanism by which this β-lactamase hydrolyzes its substrates remains poorly understood. In this study, the complex structures of OXA-48 and various β-lactams were modeled and the potential active site residues that may interact with various β-lactams were identified and characterized to elucidate their roles in OXA-48 substrate recognition. Four residues, namely S<sup>70</sup>, K<sup>73</sup>, S<sup>118</sup>, and K<sup>208</sup> were found to be essential for OXA-48 to undergo catalytic hydrolysis of various penicillins and carbapenems both in vivo and in vitro. T<sup>209</sup> was found to be important for hydrolysis of imipenem, whereas R<sup>250</sup> played a major role in hydrolyzing ampicillin, imipenem, and meropenem most likely by forming a H-bond or salt-bridge between the side chain of these two residues and the carboxylate oxygen ions of the substrates. Analysis of the effect of substitution of alanine in two residues, W<sup>105</sup> and L<sup>158</sup>, revealed their roles in mediating the activity of OXA-48. Our data show that these residues most likely undergo hydrophobic interaction with the R groups and the core structure of the β-lactam ring in penicillins and the carbapenems, respectively. Unlike OXA-58, mass spectrometry suggested a loss of the C6-hydroxyethyl group during hydrolysis of meropenem by OXA-48, which has never been demonstrated in Class D carbapenemases. Findings in this study provide comprehensive knowledge of the mechanism of the substrate recognition and catalysis of OXA-type β-lactamases.
format	article
author	Jiachi Chiou Qipeng Cheng Perry Tim-fat Shum Marcus Ho-yin Wong Edward Wai-chi Chan Sheng Chen
author_facet	Jiachi Chiou Qipeng Cheng Perry Tim-fat Shum Marcus Ho-yin Wong Edward Wai-chi Chan Sheng Chen
author_sort	Jiachi Chiou
title	Structural and Functional Characterization of OXA-48: Insight into Mechanism and Structural Basis of Substrate Recognition and Specificity
title_short	Structural and Functional Characterization of OXA-48: Insight into Mechanism and Structural Basis of Substrate Recognition and Specificity
title_full	Structural and Functional Characterization of OXA-48: Insight into Mechanism and Structural Basis of Substrate Recognition and Specificity
title_fullStr	Structural and Functional Characterization of OXA-48: Insight into Mechanism and Structural Basis of Substrate Recognition and Specificity
title_full_unstemmed	Structural and Functional Characterization of OXA-48: Insight into Mechanism and Structural Basis of Substrate Recognition and Specificity
title_sort	structural and functional characterization of oxa-48: insight into mechanism and structural basis of substrate recognition and specificity
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/f47b3272ce3a49a29acb1e096c33c58c
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Structural and Functional Characterization of OXA-48: Insight into Mechanism and Structural Basis of Substrate Recognition and Specificity

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