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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 principales: Guevara, Tibisay, Rodriguez-Banqueri, Arturo, Ksiazek, Miroslaw, Potempa, Jan, Gomis-Rüth, F. Xavier
Formato: Online Artículo Texto
Lenguaje:English
Publicado: International Union of Crystallography 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6949598/
https://www.ncbi.nlm.nih.gov/pubmed/31949901
http://dx.doi.org/10.1107/S2052252519013848
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author Guevara, Tibisay
Rodriguez-Banqueri, Arturo
Ksiazek, Miroslaw
Potempa, Jan
Gomis-Rüth, F. Xavier
author_facet Guevara, Tibisay
Rodriguez-Banqueri, Arturo
Ksiazek, Miroslaw
Potempa, Jan
Gomis-Rüth, F. Xavier
author_sort Guevara, Tibisay
collection PubMed
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 S(1)′. 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 S(1)′ to S(6)′. A citrate molecule in S(1) 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.
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spelling pubmed-69495982020-01-16 Structure-based mechanism of cysteine-switch latency and of catalysis by pappalysin-family metallopeptidases Guevara, Tibisay Rodriguez-Banqueri, Arturo Ksiazek, Miroslaw Potempa, Jan Gomis-Rüth, F. Xavier IUCrJ Research Papers 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 S(1)′. 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 S(1)′ to S(6)′. A citrate molecule in S(1) 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. International Union of Crystallography 2020-01-01 /pmc/articles/PMC6949598/ /pubmed/31949901 http://dx.doi.org/10.1107/S2052252519013848 Text en © Guevara et al. 2020 http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution (CC-BY) Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.http://creativecommons.org/licenses/by/4.0/
spellingShingle Research Papers
Guevara, Tibisay
Rodriguez-Banqueri, Arturo
Ksiazek, Miroslaw
Potempa, Jan
Gomis-Rüth, F. Xavier
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
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_short 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
topic Research Papers
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6949598/
https://www.ncbi.nlm.nih.gov/pubmed/31949901
http://dx.doi.org/10.1107/S2052252519013848
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