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PcrG protects the two long helical oligomerization domains of PcrV, by an interaction mediated by the intramolecular coiled-coil region of PcrG
BACKGROUND: PcrV is a hydrophilic translocator of type three secretion system (TTSS) and a structural component of the functional translocon. C-terminal helix of PcrV is essential for its oligomerization at the needle tip. Conformational changes within PcrV regulate the effector translocation. PcrG...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2014
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3904411/ https://www.ncbi.nlm.nih.gov/pubmed/24460624 http://dx.doi.org/10.1186/1472-6807-14-5 |
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author | Basu, Abhishek Das, Urmisha Dey, Supratim Datta, Saumen |
author_facet | Basu, Abhishek Das, Urmisha Dey, Supratim Datta, Saumen |
author_sort | Basu, Abhishek |
collection | PubMed |
description | BACKGROUND: PcrV is a hydrophilic translocator of type three secretion system (TTSS) and a structural component of the functional translocon. C-terminal helix of PcrV is essential for its oligomerization at the needle tip. Conformational changes within PcrV regulate the effector translocation. PcrG is a cytoplasmic regulator of TTSS and forms a high affinity complex with PcrV. C-terminal residues of PcrG control the effector secretion. RESULT: Both PcrV and PcrG-PcrV complex exhibit elongated conformation like their close homologs LcrV and LcrG-LcrV complex. The homology model of PcrV depicts a dumbbell shaped structure with N and C-terminal globular domains. The grip of the dumbbell is formed by two long helices (helix-7 and 12), which show high level of conservation both structurally and evolutionary. PcrG specifically protects a region of PcrV extending from helix-12 to helix-7, and encompassing the C-terminal globular domain. This fragment ∆PcrV((128–294)) interacts with PcrG with high affinity, comparable to the wild type interaction. Deletion of N-terminal globular domain leads to the oligomerization of PcrV, but PcrG restores the monomeric state of PcrV by forming a heterodimeric complex. The N-terminal globular domain (∆PcrV((1–127))) does not interact with PcrG but maintains its monomeric state. Interaction affinities of various domains of PcrV with PcrG illustrates that helix-12 is the key mediator of PcrG-PcrV interaction, supported by helix-7. Bioinformatic analysis and study with our deletion mutant ∆PcrG((13–72)) revealed that the first predicted intramolecular coiled-coil domain of PcrG contains the PcrV interaction site. However, 12 N-terminal amino acids of PcrG play an indirect role in PcrG-PcrV interaction, as their deletion causes 40-fold reduction in binding affinity and changes the kinetic parameters of interaction. ∆PcrG((13–72)) fits within the groove formed between the two globular domains of PcrV, through hydrophobic interaction. CONCLUSION: PcrG interacts with PcrV through its intramolecular coiled-coil region and masks the domains responsible for oligomerization of PcrV at the needle tip. Also, PcrG could restore the monomeric state of oligomeric PcrV. Therefore, PcrG prevents the premature oligomerization of PcrV and maintains its functional state within the bacterial cytoplasm, which is a pre-requisite for formation of the functional translocon. |
format | Online Article Text |
id | pubmed-3904411 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-39044112014-01-29 PcrG protects the two long helical oligomerization domains of PcrV, by an interaction mediated by the intramolecular coiled-coil region of PcrG Basu, Abhishek Das, Urmisha Dey, Supratim Datta, Saumen BMC Struct Biol Research Article BACKGROUND: PcrV is a hydrophilic translocator of type three secretion system (TTSS) and a structural component of the functional translocon. C-terminal helix of PcrV is essential for its oligomerization at the needle tip. Conformational changes within PcrV regulate the effector translocation. PcrG is a cytoplasmic regulator of TTSS and forms a high affinity complex with PcrV. C-terminal residues of PcrG control the effector secretion. RESULT: Both PcrV and PcrG-PcrV complex exhibit elongated conformation like their close homologs LcrV and LcrG-LcrV complex. The homology model of PcrV depicts a dumbbell shaped structure with N and C-terminal globular domains. The grip of the dumbbell is formed by two long helices (helix-7 and 12), which show high level of conservation both structurally and evolutionary. PcrG specifically protects a region of PcrV extending from helix-12 to helix-7, and encompassing the C-terminal globular domain. This fragment ∆PcrV((128–294)) interacts with PcrG with high affinity, comparable to the wild type interaction. Deletion of N-terminal globular domain leads to the oligomerization of PcrV, but PcrG restores the monomeric state of PcrV by forming a heterodimeric complex. The N-terminal globular domain (∆PcrV((1–127))) does not interact with PcrG but maintains its monomeric state. Interaction affinities of various domains of PcrV with PcrG illustrates that helix-12 is the key mediator of PcrG-PcrV interaction, supported by helix-7. Bioinformatic analysis and study with our deletion mutant ∆PcrG((13–72)) revealed that the first predicted intramolecular coiled-coil domain of PcrG contains the PcrV interaction site. However, 12 N-terminal amino acids of PcrG play an indirect role in PcrG-PcrV interaction, as their deletion causes 40-fold reduction in binding affinity and changes the kinetic parameters of interaction. ∆PcrG((13–72)) fits within the groove formed between the two globular domains of PcrV, through hydrophobic interaction. CONCLUSION: PcrG interacts with PcrV through its intramolecular coiled-coil region and masks the domains responsible for oligomerization of PcrV at the needle tip. Also, PcrG could restore the monomeric state of oligomeric PcrV. Therefore, PcrG prevents the premature oligomerization of PcrV and maintains its functional state within the bacterial cytoplasm, which is a pre-requisite for formation of the functional translocon. BioMed Central 2014-01-24 /pmc/articles/PMC3904411/ /pubmed/24460624 http://dx.doi.org/10.1186/1472-6807-14-5 Text en Copyright © 2014 Basu et al.; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Research Article Basu, Abhishek Das, Urmisha Dey, Supratim Datta, Saumen PcrG protects the two long helical oligomerization domains of PcrV, by an interaction mediated by the intramolecular coiled-coil region of PcrG |
title | PcrG protects the two long helical oligomerization domains of PcrV, by an interaction mediated by the intramolecular coiled-coil region of PcrG |
title_full | PcrG protects the two long helical oligomerization domains of PcrV, by an interaction mediated by the intramolecular coiled-coil region of PcrG |
title_fullStr | PcrG protects the two long helical oligomerization domains of PcrV, by an interaction mediated by the intramolecular coiled-coil region of PcrG |
title_full_unstemmed | PcrG protects the two long helical oligomerization domains of PcrV, by an interaction mediated by the intramolecular coiled-coil region of PcrG |
title_short | PcrG protects the two long helical oligomerization domains of PcrV, by an interaction mediated by the intramolecular coiled-coil region of PcrG |
title_sort | pcrg protects the two long helical oligomerization domains of pcrv, by an interaction mediated by the intramolecular coiled-coil region of pcrg |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3904411/ https://www.ncbi.nlm.nih.gov/pubmed/24460624 http://dx.doi.org/10.1186/1472-6807-14-5 |
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