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N-terminal-mediated oligomerization of DnaA drives the occupancy-dependent rejuvenation of the protein on the membrane

DnaA, the initiator of chromosome replication in most known eubacteria species, is activated once per cell division cycle. Its overall activity cycle is driven by ATP hydrolysis and ADP–ATP exchange. The latter can be promoted by binding to specific sequences on the chromosome and/or to acidic phosp...

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Autores principales: Aranovich, Alexander, Braier-Marcovitz, Shani, Ansbacher, Esti, Granek, Rony, Parola, Abraham H., Fishov, Itzhak
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Portland Press Ltd. 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4721551/
https://www.ncbi.nlm.nih.gov/pubmed/26272946
http://dx.doi.org/10.1042/BSR20150175
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author Aranovich, Alexander
Braier-Marcovitz, Shani
Ansbacher, Esti
Granek, Rony
Parola, Abraham H.
Fishov, Itzhak
author_facet Aranovich, Alexander
Braier-Marcovitz, Shani
Ansbacher, Esti
Granek, Rony
Parola, Abraham H.
Fishov, Itzhak
author_sort Aranovich, Alexander
collection PubMed
description DnaA, the initiator of chromosome replication in most known eubacteria species, is activated once per cell division cycle. Its overall activity cycle is driven by ATP hydrolysis and ADP–ATP exchange. The latter can be promoted by binding to specific sequences on the chromosome and/or to acidic phospholipids in the membrane. We have previously shown that the transition into an active form (rejuvenation) is strongly co-operative with respect to DnaA membrane occupancy. Only at low membrane occupancy is DnaA reactivation efficiently catalysed by the acidic phospholipids. The present study was aimed at unravelling the molecular mechanism underlying the occupancy-dependent DnaA rejuvenation. We found that truncation of the DnaA N-terminal completely abolishes the co-operative transformation between the high and low occupancy states (I and II respectively) without affecting the membrane binding. The environmentally sensitive fluorophore specifically attached to the N-terminal cysteines of DnaA reported on occupancy-correlated changes in its vicinity. Cross-linking of DnaA with a short homobifunctional reagent revealed that state II of the protein on the membrane corresponds to a distinct oligomeric form of DnaA. The kinetic transition of DnaA on the membrane surface is described in the present study by a generalized 2D condensation phase transition model, confirming the existence of two states of DnaA on the membrane and pointing to the possibility that membrane protein density serves as an on-off switch in vivo. We conclude that the DnaA conformation attained at low surface density drives its N-terminal-mediated oligomerization, which is presumably a pre-requisite for facilitated nt exchange.
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spelling pubmed-47215512016-02-02 N-terminal-mediated oligomerization of DnaA drives the occupancy-dependent rejuvenation of the protein on the membrane Aranovich, Alexander Braier-Marcovitz, Shani Ansbacher, Esti Granek, Rony Parola, Abraham H. Fishov, Itzhak Biosci Rep Original Papers DnaA, the initiator of chromosome replication in most known eubacteria species, is activated once per cell division cycle. Its overall activity cycle is driven by ATP hydrolysis and ADP–ATP exchange. The latter can be promoted by binding to specific sequences on the chromosome and/or to acidic phospholipids in the membrane. We have previously shown that the transition into an active form (rejuvenation) is strongly co-operative with respect to DnaA membrane occupancy. Only at low membrane occupancy is DnaA reactivation efficiently catalysed by the acidic phospholipids. The present study was aimed at unravelling the molecular mechanism underlying the occupancy-dependent DnaA rejuvenation. We found that truncation of the DnaA N-terminal completely abolishes the co-operative transformation between the high and low occupancy states (I and II respectively) without affecting the membrane binding. The environmentally sensitive fluorophore specifically attached to the N-terminal cysteines of DnaA reported on occupancy-correlated changes in its vicinity. Cross-linking of DnaA with a short homobifunctional reagent revealed that state II of the protein on the membrane corresponds to a distinct oligomeric form of DnaA. The kinetic transition of DnaA on the membrane surface is described in the present study by a generalized 2D condensation phase transition model, confirming the existence of two states of DnaA on the membrane and pointing to the possibility that membrane protein density serves as an on-off switch in vivo. We conclude that the DnaA conformation attained at low surface density drives its N-terminal-mediated oligomerization, which is presumably a pre-requisite for facilitated nt exchange. Portland Press Ltd. 2015-09-18 /pmc/articles/PMC4721551/ /pubmed/26272946 http://dx.doi.org/10.1042/BSR20150175 Text en © 2015 Authors http://creativecommons.org/licenses/by/3.0/ This is an open access article published by Portland Press Limited and distributed under the Creative Commons Attribution Licence 3.0 (http://creativecommons.org/licenses/by/3.0/) .
spellingShingle Original Papers
Aranovich, Alexander
Braier-Marcovitz, Shani
Ansbacher, Esti
Granek, Rony
Parola, Abraham H.
Fishov, Itzhak
N-terminal-mediated oligomerization of DnaA drives the occupancy-dependent rejuvenation of the protein on the membrane
title N-terminal-mediated oligomerization of DnaA drives the occupancy-dependent rejuvenation of the protein on the membrane
title_full N-terminal-mediated oligomerization of DnaA drives the occupancy-dependent rejuvenation of the protein on the membrane
title_fullStr N-terminal-mediated oligomerization of DnaA drives the occupancy-dependent rejuvenation of the protein on the membrane
title_full_unstemmed N-terminal-mediated oligomerization of DnaA drives the occupancy-dependent rejuvenation of the protein on the membrane
title_short N-terminal-mediated oligomerization of DnaA drives the occupancy-dependent rejuvenation of the protein on the membrane
title_sort n-terminal-mediated oligomerization of dnaa drives the occupancy-dependent rejuvenation of the protein on the membrane
topic Original Papers
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4721551/
https://www.ncbi.nlm.nih.gov/pubmed/26272946
http://dx.doi.org/10.1042/BSR20150175
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