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Antimicrob Agents Chemother. 2019 Sep 23;63(10). pii: e00957-19. doi: 10.1128/AAC.00957-19. Print 2019 Oct.

Role of MurT C-Terminal Domain in the Amidation of Staphylococcus aureus Peptidoglycan.

Author information

1
Laboratory of Molecular Microbiology of Bacterial Pathogens, UCIBIO@REQUIMTE, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Caparica, Portugal.
2
Laboratory of Molecular Genetics, Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, Oeiras, Portugal.
3
Laboratory of Microbiology and Infectious Diseases, The Rockefeller University, New York, New York, USA.
4
(Bio)Molecular Structure and Interactions by NMR, UCIBIO@REQUIMTE, Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Caparica, Portugal.
5
Laboratory of Molecular Microbiology of Bacterial Pathogens, UCIBIO@REQUIMTE, Departamento de Ciências da Vida, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Caparica, Portugal rgs@fct.unl.pt.
#
Contributed equally

Abstract

Glutamate amidation, a secondary modification of the peptidoglycan, was first identified in Staphylococcus aureus It is catalyzed by the protein products of the murT and gatD genes, which are conserved and colocalized in the genomes of most sequenced Gram-positive bacterial species. The MurT-GatD complex is required for cell viability, full resistance to β-lactam antibiotics, and resistance to human lysozyme and is recognized as an attractive target for new antimicrobials. Great effort has been invested in the study of this step, culminating recently in three independent reports addressing the structural elucidation of the MurT-GatD complex. In this work, we demonstrate through the use of nonstructural approaches the critical and multiple roles of the C-terminal domain of MurT, annotated as DUF1727, in the MurT-GatD enzymatic complex. This domain provides the physical link between the two enzymatic activities and is essential for the amidation reaction. Copurification of recombinant MurT and GatD proteins and bacterial two-hybrid assays support the observation that the MurT-GatD interaction occurs through this domain. Most importantly, we provide in vivo evidence of the effect of substitutions at specific residues in DUF1727 on cell wall peptidoglycan amidation and on the phenotypes of oxacillin resistance and bacterial growth.

KEYWORDS:

DUF1727; MurT-GatD; Staphylococcus aureus ; antibiotic resistance; cell wall; peptidoglycan amidation

PMID:
31358586
PMCID:
PMC6761511
[Available on 2020-03-23]
DOI:
10.1128/AAC.00957-19

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