Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilis
Autor(a) principal: | |
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Data de Publicação: | 2019 |
Outros Autores: | , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | https://hdl.handle.net/10216/127799 |
Resumo: | RCK (regulating conductance of K+) domains are common regulatory domains that control the activity of eukaryotic and prokaryotic K+ channels and transporters. In bacteria these domains play roles in osmoregulation, regulation of turgor and membrane potential and in pH homeostasis. Whole-genome sequencing unveiled RCK gene redundancy, however the biological role of this redundancy is not well understood. In Bacillus subtilis, there are two closely related RCK domain proteins (KtrA and KtrC) that regulate the activity of the Ktr cation channels. KtrA has been well characterized but little is known about KtrC. We have characterized the structural and biochemical proprieties of KtrC and conclude that KtrC binds ATP and ADP, just like KtrA. However, in solution KtrC exist in a dynamic equilibrium between octamers and non-octameric species that is dependent on the bound ligand, with ATP destabilizing the octameric ring relative to ADP. Accordingly, KtrC-ADP crystal structures reveal closed octameric rings similar to those in KtrA, while KtrC-ATP adopts an open assembly with RCK domains forming a super-helix. In addition, both KtrC-ATP and -ADP octamers are stabilized by the signaling molecule cyclic-di-AMP, which binds to KtrC with high affinity. In contrast, c-di-AMP binds with 100-fold lower affinity to KtrA. Despite these differences we show with an E. coli complementation assay that KtrC and KtrA are interchangeable and able to form functional transporters with both KtrB and KtrD. The distinctive properties of KtrC, in particular ligand-dependent assembly/disassembly, suggest that this protein has a specific physiological role that is distinct from KtrA. |
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Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilisAdenosine Diphosphate / chemistryAdenosine Diphosphate / metabolismAdenosine Triphosphate / chemistryAdenosine Triphosphate / metabolismAmino Acid MotifsBacillus subtilis / chemistryBacillus subtilis / metabolismBacterial Proteins / chemistryBacterial Proteins / geneticsBacterial Proteins / metabolismBinding SitesCation Transport Proteins / chemistryCation Transport Proteins / geneticsCation Transport Proteins / metabolismCations, MonovalentCloning, MolecularCrystallography, X-RayDinucleoside Phosphates / chemistryDinucleoside Phosphates / metabolismEscherichia coli / geneticsEscherichia coli / metabolismGene ExpressionGenetic Complementation TestGenetic Vectors / chemistryGenetic Vectors / metabolismIon TransportModels, MolecularPotassium / chemistryPotassium / metabolismProtein BindingProtein Conformation, alpha-HelicalProtein Conformation, beta-StrandProtein Interaction Domains and MotifsProtein Isoforms / chemistryProtein Isoforms / geneticsProtein Isoforms / metabolismProtein MultimerizationRecombinant Proteins / chemistryRecombinant Proteins / geneticsRecombinant Proteins / metabolismRCK (regulating conductance of K+) domains are common regulatory domains that control the activity of eukaryotic and prokaryotic K+ channels and transporters. In bacteria these domains play roles in osmoregulation, regulation of turgor and membrane potential and in pH homeostasis. Whole-genome sequencing unveiled RCK gene redundancy, however the biological role of this redundancy is not well understood. In Bacillus subtilis, there are two closely related RCK domain proteins (KtrA and KtrC) that regulate the activity of the Ktr cation channels. KtrA has been well characterized but little is known about KtrC. We have characterized the structural and biochemical proprieties of KtrC and conclude that KtrC binds ATP and ADP, just like KtrA. However, in solution KtrC exist in a dynamic equilibrium between octamers and non-octameric species that is dependent on the bound ligand, with ATP destabilizing the octameric ring relative to ADP. Accordingly, KtrC-ADP crystal structures reveal closed octameric rings similar to those in KtrA, while KtrC-ATP adopts an open assembly with RCK domains forming a super-helix. In addition, both KtrC-ATP and -ADP octamers are stabilized by the signaling molecule cyclic-di-AMP, which binds to KtrC with high affinity. In contrast, c-di-AMP binds with 100-fold lower affinity to KtrA. Despite these differences we show with an E. coli complementation assay that KtrC and KtrA are interchangeable and able to form functional transporters with both KtrB and KtrD. The distinctive properties of KtrC, in particular ligand-dependent assembly/disassembly, suggest that this protein has a specific physiological role that is distinct from KtrA.Academic Press20192019-01-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfapplication/vnd.openxmlformats-officedocument.wordprocessingml.documenthttps://hdl.handle.net/10216/127799eng1047-847710.1016/j.jsb.2019.02.002Rocha, RTeixeira-Duarte, CMJMP, JMorais-Cabral, JHinfo:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2023-11-29T15:41:32Zoai:repositorio-aberto.up.pt:10216/127799Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T00:29:49.244297Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse |
dc.title.none.fl_str_mv |
Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilis |
title |
Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilis |
spellingShingle |
Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilis Rocha, R Adenosine Diphosphate / chemistry Adenosine Diphosphate / metabolism Adenosine Triphosphate / chemistry Adenosine Triphosphate / metabolism Amino Acid Motifs Bacillus subtilis / chemistry Bacillus subtilis / metabolism Bacterial Proteins / chemistry Bacterial Proteins / genetics Bacterial Proteins / metabolism Binding Sites Cation Transport Proteins / chemistry Cation Transport Proteins / genetics Cation Transport Proteins / metabolism Cations, Monovalent Cloning, Molecular Crystallography, X-Ray Dinucleoside Phosphates / chemistry Dinucleoside Phosphates / metabolism Escherichia coli / genetics Escherichia coli / metabolism Gene Expression Genetic Complementation Test Genetic Vectors / chemistry Genetic Vectors / metabolism Ion Transport Models, Molecular Potassium / chemistry Potassium / metabolism Protein Binding Protein Conformation, alpha-Helical Protein Conformation, beta-Strand Protein Interaction Domains and Motifs Protein Isoforms / chemistry Protein Isoforms / genetics Protein Isoforms / metabolism Protein Multimerization Recombinant Proteins / chemistry Recombinant Proteins / genetics Recombinant Proteins / metabolism |
title_short |
Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilis |
title_full |
Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilis |
title_fullStr |
Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilis |
title_full_unstemmed |
Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilis |
title_sort |
Characterization of the molecular properties of KtrC, a second RCK domain that regulates a Ktr channel in Bacillus subtilis |
author |
Rocha, R |
author_facet |
Rocha, R Teixeira-Duarte, CM JMP, J Morais-Cabral, JH |
author_role |
author |
author2 |
Teixeira-Duarte, CM JMP, J Morais-Cabral, JH |
author2_role |
author author author |
dc.contributor.author.fl_str_mv |
Rocha, R Teixeira-Duarte, CM JMP, J Morais-Cabral, JH |
dc.subject.por.fl_str_mv |
Adenosine Diphosphate / chemistry Adenosine Diphosphate / metabolism Adenosine Triphosphate / chemistry Adenosine Triphosphate / metabolism Amino Acid Motifs Bacillus subtilis / chemistry Bacillus subtilis / metabolism Bacterial Proteins / chemistry Bacterial Proteins / genetics Bacterial Proteins / metabolism Binding Sites Cation Transport Proteins / chemistry Cation Transport Proteins / genetics Cation Transport Proteins / metabolism Cations, Monovalent Cloning, Molecular Crystallography, X-Ray Dinucleoside Phosphates / chemistry Dinucleoside Phosphates / metabolism Escherichia coli / genetics Escherichia coli / metabolism Gene Expression Genetic Complementation Test Genetic Vectors / chemistry Genetic Vectors / metabolism Ion Transport Models, Molecular Potassium / chemistry Potassium / metabolism Protein Binding Protein Conformation, alpha-Helical Protein Conformation, beta-Strand Protein Interaction Domains and Motifs Protein Isoforms / chemistry Protein Isoforms / genetics Protein Isoforms / metabolism Protein Multimerization Recombinant Proteins / chemistry Recombinant Proteins / genetics Recombinant Proteins / metabolism |
topic |
Adenosine Diphosphate / chemistry Adenosine Diphosphate / metabolism Adenosine Triphosphate / chemistry Adenosine Triphosphate / metabolism Amino Acid Motifs Bacillus subtilis / chemistry Bacillus subtilis / metabolism Bacterial Proteins / chemistry Bacterial Proteins / genetics Bacterial Proteins / metabolism Binding Sites Cation Transport Proteins / chemistry Cation Transport Proteins / genetics Cation Transport Proteins / metabolism Cations, Monovalent Cloning, Molecular Crystallography, X-Ray Dinucleoside Phosphates / chemistry Dinucleoside Phosphates / metabolism Escherichia coli / genetics Escherichia coli / metabolism Gene Expression Genetic Complementation Test Genetic Vectors / chemistry Genetic Vectors / metabolism Ion Transport Models, Molecular Potassium / chemistry Potassium / metabolism Protein Binding Protein Conformation, alpha-Helical Protein Conformation, beta-Strand Protein Interaction Domains and Motifs Protein Isoforms / chemistry Protein Isoforms / genetics Protein Isoforms / metabolism Protein Multimerization Recombinant Proteins / chemistry Recombinant Proteins / genetics Recombinant Proteins / metabolism |
description |
RCK (regulating conductance of K+) domains are common regulatory domains that control the activity of eukaryotic and prokaryotic K+ channels and transporters. In bacteria these domains play roles in osmoregulation, regulation of turgor and membrane potential and in pH homeostasis. Whole-genome sequencing unveiled RCK gene redundancy, however the biological role of this redundancy is not well understood. In Bacillus subtilis, there are two closely related RCK domain proteins (KtrA and KtrC) that regulate the activity of the Ktr cation channels. KtrA has been well characterized but little is known about KtrC. We have characterized the structural and biochemical proprieties of KtrC and conclude that KtrC binds ATP and ADP, just like KtrA. However, in solution KtrC exist in a dynamic equilibrium between octamers and non-octameric species that is dependent on the bound ligand, with ATP destabilizing the octameric ring relative to ADP. Accordingly, KtrC-ADP crystal structures reveal closed octameric rings similar to those in KtrA, while KtrC-ATP adopts an open assembly with RCK domains forming a super-helix. In addition, both KtrC-ATP and -ADP octamers are stabilized by the signaling molecule cyclic-di-AMP, which binds to KtrC with high affinity. In contrast, c-di-AMP binds with 100-fold lower affinity to KtrA. Despite these differences we show with an E. coli complementation assay that KtrC and KtrA are interchangeable and able to form functional transporters with both KtrB and KtrD. The distinctive properties of KtrC, in particular ligand-dependent assembly/disassembly, suggest that this protein has a specific physiological role that is distinct from KtrA. |
publishDate |
2019 |
dc.date.none.fl_str_mv |
2019 2019-01-01T00:00:00Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
https://hdl.handle.net/10216/127799 |
url |
https://hdl.handle.net/10216/127799 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
1047-8477 10.1016/j.jsb.2019.02.002 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf application/vnd.openxmlformats-officedocument.wordprocessingml.document |
dc.publisher.none.fl_str_mv |
Academic Press |
publisher.none.fl_str_mv |
Academic Press |
dc.source.none.fl_str_mv |
reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação instacron:RCAAP |
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Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
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RCAAP |
institution |
RCAAP |
reponame_str |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
repository.name.fl_str_mv |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
repository.mail.fl_str_mv |
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1799136208617472000 |