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Structure and mechanism of the PilF DNA transformation ATPase from Thermus thermophilus
Many Gram-negative bacteria contain specific systems for uptake of foreign DNA, which play a critical role in the acquisition of antibiotic resistance. The TtPilF (PilF ATPase from Thermus thermophilus) is required for high transformation efficiency, but its mechanism of action is unknown. In the pr...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Portland Press Ltd.
2013
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3583032/ https://www.ncbi.nlm.nih.gov/pubmed/23252471 http://dx.doi.org/10.1042/BJ20121599 |
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author | Collins, Richard F. Hassan, Darin Karuppiah, Vijaykumar Thistlethwaite, Angela Derrick, Jeremy P. |
author_facet | Collins, Richard F. Hassan, Darin Karuppiah, Vijaykumar Thistlethwaite, Angela Derrick, Jeremy P. |
author_sort | Collins, Richard F. |
collection | PubMed |
description | Many Gram-negative bacteria contain specific systems for uptake of foreign DNA, which play a critical role in the acquisition of antibiotic resistance. The TtPilF (PilF ATPase from Thermus thermophilus) is required for high transformation efficiency, but its mechanism of action is unknown. In the present study, we show that TtPilF is able to bind to both DNA and RNA. The structure of TtPilF was determined by cryoelectron microscopy in the presence and absence of the ATP analogue p[NH]ppA (adenosine 5′-[β,γ-imido]triphosphate), at 10 and 12 Å (1 Å=0.1 nm) resolutions respectively. It consists of two distinct N- and C-terminal regions, separated by a short stem-like structure. Binding of p[NH]ppA induces structural changes in the C-terminal domains, which are transmitted via the stem to the N-terminal domains. Molecular models were generated for the apoenzyme and p[NH]ppA-bound states in the C-terminal regions by docking of a model based on a crystal structure from a closely related enzyme. Analysis of DNA binding by electron microscopy, using gold labelling, localized the binding site to the N-terminal domains. The results suggest a model in which DNA uptake by TtPilF is powered by ATP hydrolysis, causing conformational changes in the C-terminal domains, which are transmitted via the stem to take up DNA into the cell. |
format | Online Article Text |
id | pubmed-3583032 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2013 |
publisher | Portland Press Ltd. |
record_format | MEDLINE/PubMed |
spelling | pubmed-35830322013-03-04 Structure and mechanism of the PilF DNA transformation ATPase from Thermus thermophilus Collins, Richard F. Hassan, Darin Karuppiah, Vijaykumar Thistlethwaite, Angela Derrick, Jeremy P. Biochem J Research Article Many Gram-negative bacteria contain specific systems for uptake of foreign DNA, which play a critical role in the acquisition of antibiotic resistance. The TtPilF (PilF ATPase from Thermus thermophilus) is required for high transformation efficiency, but its mechanism of action is unknown. In the present study, we show that TtPilF is able to bind to both DNA and RNA. The structure of TtPilF was determined by cryoelectron microscopy in the presence and absence of the ATP analogue p[NH]ppA (adenosine 5′-[β,γ-imido]triphosphate), at 10 and 12 Å (1 Å=0.1 nm) resolutions respectively. It consists of two distinct N- and C-terminal regions, separated by a short stem-like structure. Binding of p[NH]ppA induces structural changes in the C-terminal domains, which are transmitted via the stem to the N-terminal domains. Molecular models were generated for the apoenzyme and p[NH]ppA-bound states in the C-terminal regions by docking of a model based on a crystal structure from a closely related enzyme. Analysis of DNA binding by electron microscopy, using gold labelling, localized the binding site to the N-terminal domains. The results suggest a model in which DNA uptake by TtPilF is powered by ATP hydrolysis, causing conformational changes in the C-terminal domains, which are transmitted via the stem to take up DNA into the cell. Portland Press Ltd. 2013-02-15 2013-03-01 /pmc/articles/PMC3583032/ /pubmed/23252471 http://dx.doi.org/10.1042/BJ20121599 Text en © 2013 The Author(s) The author(s) has paid for this article to be freely available under the terms of the Creative Commons Attribution Non-Commercial Licence (http://creativecommons.org/licenses/by-nc/2.5/) which permits unrestricted non-commercial use, distribution and reproduction in any medium, provided the original work is properly cited. http://creativecommons.org/licenses/by-nc/2.5/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Article Collins, Richard F. Hassan, Darin Karuppiah, Vijaykumar Thistlethwaite, Angela Derrick, Jeremy P. Structure and mechanism of the PilF DNA transformation ATPase from Thermus thermophilus |
title | Structure and mechanism of the PilF DNA transformation ATPase from Thermus thermophilus |
title_full | Structure and mechanism of the PilF DNA transformation ATPase from Thermus thermophilus |
title_fullStr | Structure and mechanism of the PilF DNA transformation ATPase from Thermus thermophilus |
title_full_unstemmed | Structure and mechanism of the PilF DNA transformation ATPase from Thermus thermophilus |
title_short | Structure and mechanism of the PilF DNA transformation ATPase from Thermus thermophilus |
title_sort | structure and mechanism of the pilf dna transformation atpase from thermus thermophilus |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3583032/ https://www.ncbi.nlm.nih.gov/pubmed/23252471 http://dx.doi.org/10.1042/BJ20121599 |
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