Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases

Tannerella forsythia is an oral dysbiotic periodontopathogen involved in severe human periodontal disease. As part of its virulence factor armamentarium, at the site of colonization it secretes mirolysin, a metallopeptidase of the unicellular pappalysin family, as a zymogen that is proteolytically a...

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Autores: Guevara, Tibisay, Rodríguez-Banqueri, Arturo, Ksiazek, Miroslaw, Potempa, Jan, Gomis-Rüth, F. Xavier
Tipo de recurso: artículo
Estado:Versión publicada
Fecha de publicación:2020
País:España
Institución:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/241436
Acceso en línea:http://hdl.handle.net/10261/241436
Access Level:acceso abierto
Palabra clave:Pappalysin family
Metallo­peptidases
Mirolysin
Peridontopathogens
Zymogens
Catalytic mechanisms
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spelling Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidasesGuevara, TibisayRodríguez-Banqueri, ArturoKsiazek, MiroslawPotempa, JanGomis-Rüth, F. XavierPappalysin familyMetallo­peptidasesMirolysinPeridontopathogensZymogensCatalytic mechanismsTannerella forsythia is an oral dysbiotic periodontopathogen involved in severe human periodontal disease. As part of its virulence factor armamentarium, at the site of colonization it secretes mirolysin, a metallopeptidase of the unicellular pappalysin family, as a zymogen that is proteolytically auto-activated extracellularly at the Ser54–Arg55 bond. Crystal structures of the catalytically impaired promirolysin point mutant E225A at 1.4 and 1.6 Å revealed that latency is exerted by an N-terminal 34-residue pro-segment that shields the front surface of the 274-residue catalytic domain, thus preventing substrate access. The catalytic domain conforms to the metzincin clan of metallopeptidases and contains a double calcium site, which acts as a calcium switch for activity. The pro-segment traverses the active-site cleft in the opposite direction to the substrate, which precludes its cleavage. It is anchored to the mature enzyme through residue Arg21, which intrudes into the specificity pocket in cleft sub-site S1′. Moreover, residue Cys23 within a conserved cysteine–glycine motif blocks the catalytic zinc ion by a cysteine-switch mechanism, first described for mammalian matrix metallopeptidases. In addition, a 1.5 Å structure was obtained for a complex of mature mirolysin and a tetradecapeptide, which filled the cleft from sub-site S1′ to S6′. A citrate molecule in S1 completed a product-complex mimic that unveiled the mechanism of substrate binding and cleavage by mirolysin, the catalytic domain of which was already preformed in the zymogen. These results, including a preference for cleavage before basic residues, are likely to be valid for other unicellular pappalysins derived from archaea, bacteria, cyanobacteria, algae and fungi, including archetypal ulilysin from Methanosarcina acetivorans. They may further apply, at least in part, to the multi-domain orthologues of higher organisms.This study was supported in part by grants from Spanish, Catalan, US American (NIH/NIDR) and Polish (NCN) public agencies (BFU2015-64487R; MDM-2014-0435; Fundacio´ ‘La Marato´ de TV3’ 201815 and 2017SGR3, 2015/17/B/NZ1/ 00666, 2016/21/B/NZ1/00292, and R21DE026280). MK was recipient of a scholarship from the Polish Ministry of Science and Higher Education (1306/MOB/IV/2015/0, ‘Mobilnoc´ Plus’). The Structural Biology Unit of IBMB was a ‘Marı´a de Maeztu’ Unit of Excellence of the Spanish Ministry of Science, Innovation and Universities (2015–2019).International Union of CrystallographyGeneralitat de CatalunyaNational Institutes of Health (US)Polish Academy of SciencesMinisterio de Economía y Competitividad (España)Fundació La Marató de TV3Ministry of Science and Higher Education (Poland)Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]2021202120202021info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501Publisher's versioninfo:eu-repo/semantics/publishedVersionhttp://hdl.handle.net/10261/241436reponame:DIGITAL.CSIC. Repositorio Institucional del CSICinstname:Consejo Superior de Investigaciones Científicas (CSIC)Inglés#PLACEHOLDER_PARENT_METADATA_VALUE##PLACEHOLDER_PARENT_METADATA_VALUE#info:eu-repo/grantAgreement/MINECO/Plan Estatal de Investigación Científica y Técnica y de Innovación 2013-2016/BFU2015-64487Rinfo:eu-repo/grantAgreement/MINECO/Plan Estatal de Investigación Científica y Técnica y de Innovación 2013-2016/MDM-2014-0435http://dx.doi.org/10.1107/S2052252519013848Síinfo:eu-repo/semantics/openAccessoai:digital.csic.es:10261/2414362026-05-22T06:33:51Z
dc.title.none.fl_str_mv Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases
title Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases
spellingShingle Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases
Guevara, Tibisay
Pappalysin family
Metallo­peptidases
Mirolysin
Peridontopathogens
Zymogens
Catalytic mechanisms
title_short Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases
title_full Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases
title_fullStr Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases
title_full_unstemmed Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases
title_sort Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases
dc.creator.none.fl_str_mv Guevara, Tibisay
Rodríguez-Banqueri, Arturo
Ksiazek, Miroslaw
Potempa, Jan
Gomis-Rüth, F. Xavier
author Guevara, Tibisay
author_facet Guevara, Tibisay
Rodríguez-Banqueri, Arturo
Ksiazek, Miroslaw
Potempa, Jan
Gomis-Rüth, F. Xavier
author_role author
author2 Rodríguez-Banqueri, Arturo
Ksiazek, Miroslaw
Potempa, Jan
Gomis-Rüth, F. Xavier
author2_role author
author
author
author
dc.contributor.none.fl_str_mv Generalitat de Catalunya
National Institutes of Health (US)
Polish Academy of Sciences
Ministerio de Economía y Competitividad (España)
Fundació La Marató de TV3
Ministry of Science and Higher Education (Poland)
Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]
dc.subject.none.fl_str_mv Pappalysin family
Metallo­peptidases
Mirolysin
Peridontopathogens
Zymogens
Catalytic mechanisms
topic Pappalysin family
Metallo­peptidases
Mirolysin
Peridontopathogens
Zymogens
Catalytic mechanisms
description Tannerella forsythia is an oral dysbiotic periodontopathogen involved in severe human periodontal disease. As part of its virulence factor armamentarium, at the site of colonization it secretes mirolysin, a metallopeptidase of the unicellular pappalysin family, as a zymogen that is proteolytically auto-activated extracellularly at the Ser54–Arg55 bond. Crystal structures of the catalytically impaired promirolysin point mutant E225A at 1.4 and 1.6 Å revealed that latency is exerted by an N-terminal 34-residue pro-segment that shields the front surface of the 274-residue catalytic domain, thus preventing substrate access. The catalytic domain conforms to the metzincin clan of metallopeptidases and contains a double calcium site, which acts as a calcium switch for activity. The pro-segment traverses the active-site cleft in the opposite direction to the substrate, which precludes its cleavage. It is anchored to the mature enzyme through residue Arg21, which intrudes into the specificity pocket in cleft sub-site S1′. Moreover, residue Cys23 within a conserved cysteine–glycine motif blocks the catalytic zinc ion by a cysteine-switch mechanism, first described for mammalian matrix metallopeptidases. In addition, a 1.5 Å structure was obtained for a complex of mature mirolysin and a tetradecapeptide, which filled the cleft from sub-site S1′ to S6′. A citrate molecule in S1 completed a product-complex mimic that unveiled the mechanism of substrate binding and cleavage by mirolysin, the catalytic domain of which was already preformed in the zymogen. These results, including a preference for cleavage before basic residues, are likely to be valid for other unicellular pappalysins derived from archaea, bacteria, cyanobacteria, algae and fungi, including archetypal ulilysin from Methanosarcina acetivorans. They may further apply, at least in part, to the multi-domain orthologues of higher organisms.
publishDate 2020
dc.date.none.fl_str_mv 2020
2021
2021
2021
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
Publisher's version
info:eu-repo/semantics/publishedVersion
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/241436
url http://hdl.handle.net/10261/241436
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv #PLACEHOLDER_PARENT_METADATA_VALUE#
#PLACEHOLDER_PARENT_METADATA_VALUE#
info:eu-repo/grantAgreement/MINECO/Plan Estatal de Investigación Científica y Técnica y de Innovación 2013-2016/BFU2015-64487R
info:eu-repo/grantAgreement/MINECO/Plan Estatal de Investigación Científica y Técnica y de Innovación 2013-2016/MDM-2014-0435
http://dx.doi.org/10.1107/S2052252519013848
Sí
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.publisher.none.fl_str_mv International Union of Crystallography
publisher.none.fl_str_mv International Union of Crystallography
dc.source.none.fl_str_mv reponame:DIGITAL.CSIC. Repositorio Institucional del CSIC
instname:Consejo Superior de Investigaciones Científicas (CSIC)
instname_str Consejo Superior de Investigaciones Científicas (CSIC)
reponame_str DIGITAL.CSIC. Repositorio Institucional del CSIC
collection DIGITAL.CSIC. Repositorio Institucional del CSIC
repository.name.fl_str_mv
repository.mail.fl_str_mv
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