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VirB8 homolog TraE from plasmid pKM101 forms a hexameric ring structure and interacts with the VirB6 homolog TraD

Type IV secretion systems (T4SSs) are multiprotein assemblies that translocate macromolecules across the cell envelope of bacteria. X-ray crystallographic and electron microscopy (EM) analyses have increasingly provided structural information on individual T4SS components and on the entire complex....

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Autores principales: Casu, Bastien, Mary, Charline, Sverzhinsky, Aleksandr, Fouillen, Aurélien, Nanci, Antonio, Baron, Christian
Formato: Online Artículo Texto
Lenguaje:English
Publicado: National Academy of Sciences 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6003364/
https://www.ncbi.nlm.nih.gov/pubmed/29784815
http://dx.doi.org/10.1073/pnas.1802501115
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author Casu, Bastien
Mary, Charline
Sverzhinsky, Aleksandr
Fouillen, Aurélien
Nanci, Antonio
Baron, Christian
author_facet Casu, Bastien
Mary, Charline
Sverzhinsky, Aleksandr
Fouillen, Aurélien
Nanci, Antonio
Baron, Christian
author_sort Casu, Bastien
collection PubMed
description Type IV secretion systems (T4SSs) are multiprotein assemblies that translocate macromolecules across the cell envelope of bacteria. X-ray crystallographic and electron microscopy (EM) analyses have increasingly provided structural information on individual T4SS components and on the entire complex. As of now, relatively little information has been available on the exact localization of the inner membrane-bound T4SS components, notably the mostly periplasmic VirB8 protein and the very hydrophobic VirB6 protein. We show here that the membrane-bound, full-length version of the VirB8 homolog TraE from the plasmid pKM101 secretion system forms a high-molecular-mass complex that is distinct from the previously characterized periplasmic portion of the protein that forms dimers. Full-length TraE was extracted from the membranes with detergents, and analysis by size-exclusion chromatography, cross-linking, and size exclusion chromatography (SEC) multiangle light scattering (MALS) shows that it forms a high-molecular-mass complex. EM and small-angle X-ray scattering (SAXS) analysis demonstrate that full-length TraE forms a hexameric complex with a central pore. We also overproduced and purified the VirB6 homolog TraD and show by cross-linking, SEC, and EM that it binds to TraE. Our results suggest that TraE and TraD interact at the substrate translocation pore of the secretion system.
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spelling pubmed-60033642018-06-18 VirB8 homolog TraE from plasmid pKM101 forms a hexameric ring structure and interacts with the VirB6 homolog TraD Casu, Bastien Mary, Charline Sverzhinsky, Aleksandr Fouillen, Aurélien Nanci, Antonio Baron, Christian Proc Natl Acad Sci U S A Biological Sciences Type IV secretion systems (T4SSs) are multiprotein assemblies that translocate macromolecules across the cell envelope of bacteria. X-ray crystallographic and electron microscopy (EM) analyses have increasingly provided structural information on individual T4SS components and on the entire complex. As of now, relatively little information has been available on the exact localization of the inner membrane-bound T4SS components, notably the mostly periplasmic VirB8 protein and the very hydrophobic VirB6 protein. We show here that the membrane-bound, full-length version of the VirB8 homolog TraE from the plasmid pKM101 secretion system forms a high-molecular-mass complex that is distinct from the previously characterized periplasmic portion of the protein that forms dimers. Full-length TraE was extracted from the membranes with detergents, and analysis by size-exclusion chromatography, cross-linking, and size exclusion chromatography (SEC) multiangle light scattering (MALS) shows that it forms a high-molecular-mass complex. EM and small-angle X-ray scattering (SAXS) analysis demonstrate that full-length TraE forms a hexameric complex with a central pore. We also overproduced and purified the VirB6 homolog TraD and show by cross-linking, SEC, and EM that it binds to TraE. Our results suggest that TraE and TraD interact at the substrate translocation pore of the secretion system. National Academy of Sciences 2018-06-05 2018-05-21 /pmc/articles/PMC6003364/ /pubmed/29784815 http://dx.doi.org/10.1073/pnas.1802501115 Text en Copyright © 2018 the Author(s). Published by PNAS. https://creativecommons.org/licenses/by-nc-nd/4.0/ This open access article is distributed under Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND) (https://creativecommons.org/licenses/by-nc-nd/4.0/) .
spellingShingle Biological Sciences
Casu, Bastien
Mary, Charline
Sverzhinsky, Aleksandr
Fouillen, Aurélien
Nanci, Antonio
Baron, Christian
VirB8 homolog TraE from plasmid pKM101 forms a hexameric ring structure and interacts with the VirB6 homolog TraD
title VirB8 homolog TraE from plasmid pKM101 forms a hexameric ring structure and interacts with the VirB6 homolog TraD
title_full VirB8 homolog TraE from plasmid pKM101 forms a hexameric ring structure and interacts with the VirB6 homolog TraD
title_fullStr VirB8 homolog TraE from plasmid pKM101 forms a hexameric ring structure and interacts with the VirB6 homolog TraD
title_full_unstemmed VirB8 homolog TraE from plasmid pKM101 forms a hexameric ring structure and interacts with the VirB6 homolog TraD
title_short VirB8 homolog TraE from plasmid pKM101 forms a hexameric ring structure and interacts with the VirB6 homolog TraD
title_sort virb8 homolog trae from plasmid pkm101 forms a hexameric ring structure and interacts with the virb6 homolog trad
topic Biological Sciences
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6003364/
https://www.ncbi.nlm.nih.gov/pubmed/29784815
http://dx.doi.org/10.1073/pnas.1802501115
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