Structural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases

ABSTRACT Vancomycin resistance in Gram-positive bacteria results from the replacement of the d-alanyl–d-alanine target of peptidoglycan precursors with d-alanyl–d-lactate or d-alanyl–d-serine (d-Ala-d-Ser), to which vancomycin has low binding affinity. VanT is one of the proteins required for the pr...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Djalal Meziane-Cherif, Peter J. Stogios, Elena Evdokimova, Olga Egorova, Alexei Savchenko, Patrice Courvalin
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2015
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/66fcbe5854e04991a978976db2deb68a
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:66fcbe5854e04991a978976db2deb68a
record_format	dspace
spelling	oai:doaj.org-article:66fcbe5854e04991a978976db2deb68a2021-11-15T15:41:26ZStructural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases10.1128/mBio.00806-152150-7511https://doaj.org/article/66fcbe5854e04991a978976db2deb68a2015-09-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00806-15https://doaj.org/toc/2150-7511ABSTRACT Vancomycin resistance in Gram-positive bacteria results from the replacement of the d-alanyl–d-alanine target of peptidoglycan precursors with d-alanyl–d-lactate or d-alanyl–d-serine (d-Ala-d-Ser), to which vancomycin has low binding affinity. VanT is one of the proteins required for the production of d-Ala-d-Ser-terminating precursors by converting l-Ser to d-Ser. VanT is composed of two domains, an N-terminal membrane-bound domain, likely involved in l-Ser uptake, and a C-terminal cytoplasmic catalytic domain which is related to bacterial alanine racemases. To gain insight into the molecular function of VanT, the crystal structure of the catalytic domain of VanTG from VanG-type resistant Enterococcus faecalis BM4518 was determined. The structure showed significant similarity to type III pyridoxal 5′-phosphate (PLP)-dependent alanine racemases, which are essential for peptidoglycan synthesis. Comparative structural analysis between VanTG and alanine racemases as well as site-directed mutagenesis identified three specific active site positions centered around Asn696 which are responsible for the l-amino acid specificity. This analysis also suggested that VanT racemases evolved from regular alanine racemases by acquiring additional selectivity toward serine while preserving that for alanine. The 4-fold-lower relative catalytic efficiency of VanTG against l-Ser versus l-Ala implied that this enzyme relies on its membrane-bound domain for l-Ser transport to increase the overall rate of d-Ser production. These findings illustrate how vancomycin pressure selected for molecular adaptation of a housekeeping enzyme to a bifunctional enzyme to allow for peptidoglycan remodeling, a strategy increasingly observed in antibiotic-resistant bacteria. IMPORTANCE Vancomycin is one of the drugs of last resort against Gram-positive antibiotic-resistant pathogens. However, bacteria have evolved a sophisticated mechanism which remodels the drug target, the d-alanine ending precursors in cell wall synthesis, into precursors terminating with d-lactate or d-serine, to which vancomycin has less affinity. d-Ser is synthesized by VanT serine racemase, which has two unusual characteristics: (i) it is one of the few serine racemases identified in bacteria and (ii) it contains a membrane-bound domain involved in l-Ser uptake. The structure of the catalytic domain of VanTG showed high similarity to alanine racemases, and we identified three specific active site substitutions responsible for l-Ser specificity. The data provide the molecular basis for VanT evolution to a bifunctional enzyme coordinating both transport and racemization. Our findings also illustrate the evolution of the essential alanine racemase into a vancomycin resistance enzyme in response to antibiotic pressure.Djalal Meziane-CherifPeter J. StogiosElena EvdokimovaOlga EgorovaAlexei SavchenkoPatrice CourvalinAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 6, Iss 4 (2015)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Djalal Meziane-Cherif Peter J. Stogios Elena Evdokimova Olga Egorova Alexei Savchenko Patrice Courvalin Structural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases
description	ABSTRACT Vancomycin resistance in Gram-positive bacteria results from the replacement of the d-alanyl–d-alanine target of peptidoglycan precursors with d-alanyl–d-lactate or d-alanyl–d-serine (d-Ala-d-Ser), to which vancomycin has low binding affinity. VanT is one of the proteins required for the production of d-Ala-d-Ser-terminating precursors by converting l-Ser to d-Ser. VanT is composed of two domains, an N-terminal membrane-bound domain, likely involved in l-Ser uptake, and a C-terminal cytoplasmic catalytic domain which is related to bacterial alanine racemases. To gain insight into the molecular function of VanT, the crystal structure of the catalytic domain of VanTG from VanG-type resistant Enterococcus faecalis BM4518 was determined. The structure showed significant similarity to type III pyridoxal 5′-phosphate (PLP)-dependent alanine racemases, which are essential for peptidoglycan synthesis. Comparative structural analysis between VanTG and alanine racemases as well as site-directed mutagenesis identified three specific active site positions centered around Asn696 which are responsible for the l-amino acid specificity. This analysis also suggested that VanT racemases evolved from regular alanine racemases by acquiring additional selectivity toward serine while preserving that for alanine. The 4-fold-lower relative catalytic efficiency of VanTG against l-Ser versus l-Ala implied that this enzyme relies on its membrane-bound domain for l-Ser transport to increase the overall rate of d-Ser production. These findings illustrate how vancomycin pressure selected for molecular adaptation of a housekeeping enzyme to a bifunctional enzyme to allow for peptidoglycan remodeling, a strategy increasingly observed in antibiotic-resistant bacteria. IMPORTANCE Vancomycin is one of the drugs of last resort against Gram-positive antibiotic-resistant pathogens. However, bacteria have evolved a sophisticated mechanism which remodels the drug target, the d-alanine ending precursors in cell wall synthesis, into precursors terminating with d-lactate or d-serine, to which vancomycin has less affinity. d-Ser is synthesized by VanT serine racemase, which has two unusual characteristics: (i) it is one of the few serine racemases identified in bacteria and (ii) it contains a membrane-bound domain involved in l-Ser uptake. The structure of the catalytic domain of VanTG showed high similarity to alanine racemases, and we identified three specific active site substitutions responsible for l-Ser specificity. The data provide the molecular basis for VanT evolution to a bifunctional enzyme coordinating both transport and racemization. Our findings also illustrate the evolution of the essential alanine racemase into a vancomycin resistance enzyme in response to antibiotic pressure.
format	article
author	Djalal Meziane-Cherif Peter J. Stogios Elena Evdokimova Olga Egorova Alexei Savchenko Patrice Courvalin
author_facet	Djalal Meziane-Cherif Peter J. Stogios Elena Evdokimova Olga Egorova Alexei Savchenko Patrice Courvalin
author_sort	Djalal Meziane-Cherif
title	Structural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases
title_short	Structural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases
title_full	Structural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases
title_fullStr	Structural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases
title_full_unstemmed	Structural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases
title_sort	structural and functional adaptation of vancomycin resistance vant serine racemases
publisher	American Society for Microbiology
publishDate	2015
url	https://doaj.org/article/66fcbe5854e04991a978976db2deb68a
work_keys_str_mv	AT djalalmezianecherif structuralandfunctionaladaptationofvancomycinresistancevantserineracemases AT peterjstogios structuralandfunctionaladaptationofvancomycinresistancevantserineracemases AT elenaevdokimova structuralandfunctionaladaptationofvancomycinresistancevantserineracemases AT olgaegorova structuralandfunctionaladaptationofvancomycinresistancevantserineracemases AT alexeisavchenko structuralandfunctionaladaptationofvancomycinresistancevantserineracemases AT patricecourvalin structuralandfunctionaladaptationofvancomycinresistancevantserineracemases
_version_	1718427708410560512

Structural and Functional Adaptation of Vancomycin Resistance VanT Serine Racemases

Ejemplares similares