A Unique Sequence Is Essential for Efficient Multidrug Efflux Function of the MtrD Protein of <italic toggle="yes">Neisseria gonorrhoeae</italic>

ABSTRACT Antimicrobial resistance in Neisseria gonorrhoeae has reached an alarming level, severely impacting the effective treatment of gonorrhea. Belonging to the resistance-nodulation-cell division (RND) superfamily of efflux transporters, the MtrD membrane protein of N. gonorrhoeae provides resis...

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Autores principales:	Mohsen Chitsaz, Vrinda Gupta, Benjamin Harris, Megan L. O’Mara, Melissa H. Brown
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2021
Materias:	RND protein efflux pump Neisseria gonorrhoeae gonorrhea multidrug resistance MtrD protein Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/08dd0a5af4dd47118014eb303e4ea2de
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spelling	oai:doaj.org-article:08dd0a5af4dd47118014eb303e4ea2de2021-11-10T18:37:52ZA Unique Sequence Is Essential for Efficient Multidrug Efflux Function of the MtrD Protein of <italic toggle="yes">Neisseria gonorrhoeae</italic>10.1128/mBio.01675-212150-7511https://doaj.org/article/08dd0a5af4dd47118014eb303e4ea2de2021-08-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01675-21https://doaj.org/toc/2150-7511ABSTRACT Antimicrobial resistance in Neisseria gonorrhoeae has reached an alarming level, severely impacting the effective treatment of gonorrhea. Belonging to the resistance-nodulation-cell division (RND) superfamily of efflux transporters, the MtrD membrane protein of N. gonorrhoeae provides resistance to a broad range of antimicrobial compounds. A unique feature of MtrD is an 11-residue sequence (from N917 to P927 [N917-P927]) that connects transmembrane helices (TMS) 9 and 10; this sequence is not present in homologous RND proteins. This study explores the structural and functional roles of the N917-P927 region by means of mutant analysis and molecular dynamics simulations. We show that N917-P927 plays a key role in modulating substrate access to the binding cleft and influences the overall orientation of the protein within the inner membrane necessary for optimal functioning. Removal of N917-P927 significantly reduced MtrD-mediated resistance to a range of antimicrobials and mutations of three single amino acids impacted MtrD-mediated multidrug resistance. Furthermore, molecular dynamics simulations showed deletion of N917-P927 in MtrD may dysregulate access of the substrate to the binding cleft and closure of the substrate-binding pocket during the transport cycle. These findings indicate that N917-P927 is a key region for interacting with the inner membrane, conceivably influencing substrate capture from the membrane-periplasm interface and thus is essential for full multidrug resistance capacity of MtrD. IMPORTANCE The historical sexually transmitted infection gonorrhea continues to be a major public health concern with an estimated global annual incidence of 86.9 million cases. N. gonorrhoeae has been identified by the World Health Organization as one of the 12 antimicrobial-resistant bacterial species that poses the greatest risk to human health. As the major efflux pump in gonococci, the MtrD transporter contributes to the cell envelope barrier in this organism and pumps antimicrobials from the periplasm and inner membrane, resulting in resistance. This study demonstrates that a unique region of the MtrD protein that connects TMS 9 and TMS 10 forms a structure that may interact with the inner membrane positioning TMS 9 and stabilizing the protein facilitating substrate capture from the inner membrane-periplasm interface. Analysis of mutants of this region identified that it was essential for MtrD-mediated multidrug resistance. Characterization of the structure and function of this unique local region of MtrD has implications for drug efflux mechanisms used by related proteins and is important knowledge for development of antibiotics that bypass efflux.Mohsen ChitsazVrinda GuptaBenjamin HarrisMegan L. O’MaraMelissa H. BrownAmerican Society for MicrobiologyarticleRND proteinefflux pumpNeisseria gonorrhoeaegonorrheamultidrug resistanceMtrD proteinMicrobiologyQR1-502ENmBio, Vol 12, Iss 4 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	RND protein efflux pump Neisseria gonorrhoeae gonorrhea multidrug resistance MtrD protein Microbiology QR1-502
spellingShingle	RND protein efflux pump Neisseria gonorrhoeae gonorrhea multidrug resistance MtrD protein Microbiology QR1-502 Mohsen Chitsaz Vrinda Gupta Benjamin Harris Megan L. O’Mara Melissa H. Brown A Unique Sequence Is Essential for Efficient Multidrug Efflux Function of the MtrD Protein of <italic toggle="yes">Neisseria gonorrhoeae</italic>
description	ABSTRACT Antimicrobial resistance in Neisseria gonorrhoeae has reached an alarming level, severely impacting the effective treatment of gonorrhea. Belonging to the resistance-nodulation-cell division (RND) superfamily of efflux transporters, the MtrD membrane protein of N. gonorrhoeae provides resistance to a broad range of antimicrobial compounds. A unique feature of MtrD is an 11-residue sequence (from N917 to P927 [N917-P927]) that connects transmembrane helices (TMS) 9 and 10; this sequence is not present in homologous RND proteins. This study explores the structural and functional roles of the N917-P927 region by means of mutant analysis and molecular dynamics simulations. We show that N917-P927 plays a key role in modulating substrate access to the binding cleft and influences the overall orientation of the protein within the inner membrane necessary for optimal functioning. Removal of N917-P927 significantly reduced MtrD-mediated resistance to a range of antimicrobials and mutations of three single amino acids impacted MtrD-mediated multidrug resistance. Furthermore, molecular dynamics simulations showed deletion of N917-P927 in MtrD may dysregulate access of the substrate to the binding cleft and closure of the substrate-binding pocket during the transport cycle. These findings indicate that N917-P927 is a key region for interacting with the inner membrane, conceivably influencing substrate capture from the membrane-periplasm interface and thus is essential for full multidrug resistance capacity of MtrD. IMPORTANCE The historical sexually transmitted infection gonorrhea continues to be a major public health concern with an estimated global annual incidence of 86.9 million cases. N. gonorrhoeae has been identified by the World Health Organization as one of the 12 antimicrobial-resistant bacterial species that poses the greatest risk to human health. As the major efflux pump in gonococci, the MtrD transporter contributes to the cell envelope barrier in this organism and pumps antimicrobials from the periplasm and inner membrane, resulting in resistance. This study demonstrates that a unique region of the MtrD protein that connects TMS 9 and TMS 10 forms a structure that may interact with the inner membrane positioning TMS 9 and stabilizing the protein facilitating substrate capture from the inner membrane-periplasm interface. Analysis of mutants of this region identified that it was essential for MtrD-mediated multidrug resistance. Characterization of the structure and function of this unique local region of MtrD has implications for drug efflux mechanisms used by related proteins and is important knowledge for development of antibiotics that bypass efflux.
format	article
author	Mohsen Chitsaz Vrinda Gupta Benjamin Harris Megan L. O’Mara Melissa H. Brown
author_facet	Mohsen Chitsaz Vrinda Gupta Benjamin Harris Megan L. O’Mara Melissa H. Brown
author_sort	Mohsen Chitsaz
title	A Unique Sequence Is Essential for Efficient Multidrug Efflux Function of the MtrD Protein of <italic toggle="yes">Neisseria gonorrhoeae</italic>
title_short	A Unique Sequence Is Essential for Efficient Multidrug Efflux Function of the MtrD Protein of <italic toggle="yes">Neisseria gonorrhoeae</italic>
title_full	A Unique Sequence Is Essential for Efficient Multidrug Efflux Function of the MtrD Protein of <italic toggle="yes">Neisseria gonorrhoeae</italic>
title_fullStr	A Unique Sequence Is Essential for Efficient Multidrug Efflux Function of the MtrD Protein of <italic toggle="yes">Neisseria gonorrhoeae</italic>
title_full_unstemmed	A Unique Sequence Is Essential for Efficient Multidrug Efflux Function of the MtrD Protein of <italic toggle="yes">Neisseria gonorrhoeae</italic>
title_sort	unique sequence is essential for efficient multidrug efflux function of the mtrd protein of <italic toggle="yes">neisseria gonorrhoeae</italic>
publisher	American Society for Microbiology
publishDate	2021
url	https://doaj.org/article/08dd0a5af4dd47118014eb303e4ea2de
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A Unique Sequence Is Essential for Efficient Multidrug Efflux Function of the MtrD Protein of <italic toggle="yes">Neisseria gonorrhoeae</italic>

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