Logo do repositório
 
Publicação

Molecular Dissection of Escherichia coli CpdB: Roles of the N Domain in Catalysis and Phosphate Inhibition, and of the C Domain in Substrate Specificity and Adenosine Inhibition

2021-02-17Artigo científico Acesso aberto
Ver registo simples
datacite.subject.fosCiências Naturais::Ciências Químicas
datacite.subject.fosCiências Naturais::Ciências da Computação e da Informação
datacite.subject.fosCiências Naturais::Ciências Biológicas
datacite.subject.fosEngenharia e Tecnologia::Engenharia Química
datacite.subject.sdg06:Água Potável e Saneamento
datacite.subject.sdg11:Cidades e Comunidades Sustentáveis
datacite.subject.sdg13:Ação Climática
dc.contributor.authorLópez-Villamizar, Iralis
dc.contributor.authorCabezas, Alicia
dc.contributor.authorPinto, Rosa María
dc.contributor.authorCanales, José
dc.contributor.authorRibeiro, João Meireles
dc.contributor.authorRodrigues, Joaquim Rui
dc.contributor.authorCostas, María Jesús
dc.contributor.authorCameselle, José Carlos
dc.date.accessioned2026-03-30T14:14:54Z
dc.date.available2026-03-30T14:14:54Z
dc.date.issued2021-02-17
dc.description.abstractCpdB is a 3′-nucleotidase/2′ 3′-cyclic nucleotide phosphodiesterase, active also with rea-sonable efficiency on cyclic dinucleotides like c-di-AMP (3′,5′-cyclic diadenosine monophosphate) and c-di-GMP (3′,5′-cyclic diadenosine monophosphate). These are regulators of bacterial physi-ology, but are also pathogen-associated molecular patterns recognized by STING to induce IFN-β response in infected hosts. The cpdB gene of Gram-negative and its homologs of gram-positive bacteria are virulence factors. Their protein products are extracytoplasmic enzymes (either periplas-mic or cell–wall anchored) and can hydrolyze extracellular cyclic dinucleotides, thus reducing the innate immune responses of infected hosts. This makes CpdB(-like) enzymes potential targets for novel therapeutic strategies in infectious diseases, bringing about the necessity to gain insight into the molecular bases of their catalytic behavior. We have dissected the two-domain structure of Escherichia coli CpdB to study the role of its N-terminal and C-terminal domains (CpdB_Ndom and CpdB_Cdom). The specificity, kinetics and inhibitor sensitivity of point mutants of CpdB, and truncated proteins CpdB_Ndom and CpdB_Cdom were investigated. CpdB_Ndom contains the catalytic site, is inhibited by phosphate but not by adenosine, while CpdB_Cdom is inactive but contains a substrate-binding site that determines substrate specificity and adenosine inhibition of CpdB. Among CpdB substrates, 3′-AMP, cyclic dinucleotides and linear dinucleotides are strongly dependent on the CpdB_Cdom binding site for activity, as the isolated CpdB_Ndom showed much-diminished activity on them. In contrast, 2′,3′-cyclic mononucleotides and bis-4-nitrophenylphosphate were actively hydrolyzed by CpdB_Ndom, indicating that they are rather independent of the CpdB_Cdom binding site.eng
dc.description.sponsorshipThis research was funded by Consejería de Economía, Ciencia y Agenda Digital, Junta de Extremadura (grant numbers IB16066 and GR18127), co-funded by FEDER (European Regional Development Fund). The APC was funded by grant GR18127. The work in Leiria (J.R.R.) was financially supported by Associate Laboratory LSRE-LCM-Base Funding UIDB/50020/2020, funded by national funds through FCT/MCTES (PIDDAC).
dc.identifier.citationLópez-Villamizar, I.; Cabezas, A.; Pinto, R.M.; Canales, J.; Ribeiro, J.M.; Rodrigues, J.R.; Costas, M.J.; Cameselle, J.C. Molecular Dissection of Escherichia coli CpdB: Roles of the N Domain in Catalysis and Phosphate Inhibition, and of the C Domain in Substrate Specificity and Adenosine Inhibition. Int. J. Mol. Sci. 2021, 22, 1977. https://doi.org/10.3390/ijms22041977.
dc.identifier.doi10.3390/ijms22041977
dc.identifier.eissn1422-0067
dc.identifier.issn1661-6596
dc.identifier.urihttp://hdl.handle.net/10400.8/16053
dc.language.isoeng
dc.peerreviewedyes
dc.publisherMDPI
dc.relationLaboratory of Separation and Reaction Engineering - Laboratory of Catalysis and Materials
dc.relation.hasversionhttps://www.mdpi.com/1422-0067/22/4/1977
dc.relation.ispartofInternational Journal of Molecular Sciences
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/
dc.subjectpathogen–host interaction
dc.subjectcyclic dinucleotide
dc.subjectextracytoplasmic phosphodiesterase
dc.subjectprotein domain
dc.subjecttruncated protein
dc.subjectpoint mutant
dc.subjectsubstrate-binding site
dc.subjectcatalytic site
dc.subjectsubstrate specificity
dc.subjectinhibitor sensitivity
dc.titleMolecular Dissection of Escherichia coli CpdB: Roles of the N Domain in Catalysis and Phosphate Inhibition, and of the C Domain in Substrate Specificity and Adenosine Inhibitioneng
dc.typejournal article
dcterms.referenceshttps://www.mdpi.com/1422-0067/22/4/1977/s1
dspace.entity.typePublication
oaire.awardNumberUIDB/50020/2020
oaire.awardTitleLaboratory of Separation and Reaction Engineering - Laboratory of Catalysis and Materials
oaire.awardURIinfo:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F50020%2F2020/PT
oaire.citation.endPage18
oaire.citation.issue4
oaire.citation.startPage1
oaire.citation.titleInternational Journal of Molecular Sciences
oaire.citation.volume22
oaire.fundingStream6817 - DCRRNI ID
oaire.versionhttp://purl.org/coar/version/c_970fb48d4fbd8a85
person.familyNameRodrigues
person.givenNameJoaquim Rui
person.identifier.ciencia-id9018-0B83-E2C6
person.identifier.orcid0000-0002-9756-1124
person.identifier.ridL-4137-2014
person.identifier.scopus-author-id10242931100
project.funder.identifierhttp://doi.org/10.13039/501100001871
project.funder.nameFundação para a Ciência e a Tecnologia
relation.isAuthorOfPublication52f6ffb2-43e9-4f78-b414-dbac46f80305
relation.isAuthorOfPublication.latestForDiscovery52f6ffb2-43e9-4f78-b414-dbac46f80305
relation.isProjectOfPublicationb8cc6c4f-3181-4666-bd4e-ed993f8cd85d
relation.isProjectOfPublication.latestForDiscoveryb8cc6c4f-3181-4666-bd4e-ed993f8cd85d

Ficheiros

Principais
A mostrar 1 - 1 de 1
Miniatura
Nome:
Article molecular dissection of escherichia coli cpdb Roles of the n domain in catalysis and phosphate inhibition, and of the c domain in substrate specificity an.pdf
Tamanho:
4.33 MB
Formato:
Adobe Portable Document Format
Descrição:
CpdB is a 3′-nucleotidase/2′ 3′-cyclic nucleotide phosphodiesterase, active also with rea-sonable efficiency on cyclic dinucleotides like c-di-AMP (3′,5′-cyclic diadenosine monophosphate) and c-di-GMP (3′,5′-cyclic diadenosine monophosphate). These are regulators of bacterial physi-ology, but are also pathogen-associated molecular patterns recognized by STING to induce IFN-β response in infected hosts. The cpdB gene of Gram-negative and its homologs of gram-positive bacteria are virulence factors. Their protein products are extracytoplasmic enzymes (either periplas-mic or cell–wall anchored) and can hydrolyze extracellular cyclic dinucleotides, thus reducing the innate immune responses of infected hosts. This makes CpdB(-like) enzymes potential targets for novel therapeutic strategies in infectious diseases, bringing about the necessity to gain insight into the molecular bases of their catalytic behavior. We have dissected the two-domain structure of Escherichia coli CpdB to study the role of its N-terminal and C-terminal domains (CpdB_Ndom and CpdB_Cdom). The specificity, kinetics and inhibitor sensitivity of point mutants of CpdB, and truncated proteins CpdB_Ndom and CpdB_Cdom were investigated. CpdB_Ndom contains the catalytic site, is inhibited by phosphate but not by adenosine, while CpdB_Cdom is inactive but contains a substrate-binding site that determines substrate specificity and adenosine inhibition of CpdB. Among CpdB substrates, 3′-AMP, cyclic dinucleotides and linear dinucleotides are strongly dependent on the CpdB_Cdom binding site for activity, as the isolated CpdB_Ndom showed much-diminished activity on them. In contrast, 2′,3′-cyclic mononucleotides and bis-4-nitrophenylphosphate were actively hydrolyzed by CpdB_Ndom, indicating that they are rather independent of the CpdB_Cdom binding site.
Ver/Abrir
Licença
A mostrar 1 - 1 de 1
Miniatura indisponível
Nome:
license.txt
Tamanho:
1.32 KB
Formato:
Item-specific license agreed upon to submission
Descrição:
Ver/Abrir

Coleções

LSRE-LCM - Artigos em Revistas Nacionais e de Circulação Internacional
ESTG - Artigos em revistas internacionais