Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracasei

The linear polymer polyphosphate (poly-P) is present across all three domains of life and serves diverse physiological functions. The enzyme polyphosphate kinase (Ppk) is responsible for poly-P synthesis, whereas poly-P degradation is carried out by the enzyme exopolyphosphatase (Ppx). In many Lacto...

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Autores: Corrales, Daniela, Alcántara, Cristina, Zúñiga, Manuel, Monedero, Vicente
Tipo de recurso: artículo
Estado:Versión aceptada para publicación
Fecha de publicación:2024
País:España
Institución:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/355737
Acceso en línea:http://hdl.handle.net/10261/355737
Access Level:acceso abierto
Palabra clave:Lacticaseibacillus paracasei
Exopolyphosphatase
Polyphosphate
Polyphosphate kinase
lactic acid bacteria
polyphosphates
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network_name_str España
repository_id_str
dc.title.none.fl_str_mv Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracasei
title Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracasei
spellingShingle Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracasei
Corrales, Daniela
Lacticaseibacillus paracasei
Exopolyphosphatase
Polyphosphate
Polyphosphate kinase
lactic acid bacteria
polyphosphates
title_short Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracasei
title_full Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracasei
title_fullStr Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracasei
title_full_unstemmed Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracasei
title_sort Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracasei
dc.creator.none.fl_str_mv Corrales, Daniela
Alcántara, Cristina
Zúñiga, Manuel
Monedero, Vicente
author Corrales, Daniela
author_facet Corrales, Daniela
Alcántara, Cristina
Zúñiga, Manuel
Monedero, Vicente
author_role author
author2 Alcántara, Cristina
Zúñiga, Manuel
Monedero, Vicente
author2_role author
author
author
dc.contributor.none.fl_str_mv Ministerio de Ciencia, Tecnología e Innovación (Colombia)
Agencia Estatal de Investigación (España)
European Commission
Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]
dc.subject.none.fl_str_mv Lacticaseibacillus paracasei
Exopolyphosphatase
Polyphosphate
Polyphosphate kinase
lactic acid bacteria
polyphosphates
topic Lacticaseibacillus paracasei
Exopolyphosphatase
Polyphosphate
Polyphosphate kinase
lactic acid bacteria
polyphosphates
description The linear polymer polyphosphate (poly-P) is present across all three domains of life and serves diverse physiological functions. The enzyme polyphosphate kinase (Ppk) is responsible for poly-P synthesis, whereas poly-P degradation is carried out by the enzyme exopolyphosphatase (Ppx). In many Lactobacillaceae, the Ppk-encoding gene (ppk) is found clustered together with two genes encoding putative exopolyphosphatases (ppx1 and ppx2) each having different domain compositions, with the gene order ppx1-ppk-ppx2. However, the specific function of these ppx genes remains unexplored. An in-frame deletion of ppx1 in Lacticaseibacillus paracasei BL23 resulted in bacteria unable to accumulate poly-P, whereas the disruption of ppx2 did not affect poly-P synthesis. The expression of ppk was not altered in the Δppx1 strain, and poly-P synthesis in this strain was only restored by expressing ppx1 in trans. Moreover, no poly-P synthesis was observed when ppk was expressed from a plasmid in the Δppx1 strain. Purified Ppx2 exhibited in vitro exopolyphosphatase activity, whereas no in vitro enzymatic activity could be demonstrated for Ppx1. This observation corresponds with the absence in Ppx1 of conserved motifs essential for catalysis found in characterized exopolyphosphatases. Furthermore, assays with purified Ppk and Ppx1 evidenced that Ppx1 enhanced Ppk activity. These results demonstrate that Ppx1 is essential for poly-P synthesis in Lc. paracasei and have unveiled, for the first time, an unexpected role of Ppx1 exopolyphosphatase in poly-P synthesis.IMPORTANCEPoly-P is a pivotal molecular player in bacteria, participating in a diverse array of processes ranging from stress resilience to pathogenesis while also serving as a functional component in probiotic bacteria. The synthesis of poly-P is tightly regulated, but the underlying mechanisms remain incompletely elucidated. Our study sheds light on the distinctive role played by the two exopolyphosphatases (Ppx) found in the Lactobacillaceae bacterial group, of relevance in food and health. This particular group is noteworthy for possessing two Ppx enzymes, supposedly involved in poly-P degradation. Remarkably, our investigation uncovers an unprecedented function of Ppx1 in Lacticaseibacillus paracasei, where its absence leads to the total cessation of poly-P synthesis, paralleling the impact observed upon eliminating the poly-P forming enzyme, poly-P kinase. Unlike the anticipated role as a conventional exopolyphosphatase, Ppx1 demonstrates an unexpected function. Our results added a layer of complexity to our understanding of poly-P dynamics in bacteria.
publishDate 2024
dc.date.none.fl_str_mv 2024
2024
2024
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
Postprint
info:eu-repo/semantics/acceptedVersion
format article
status_str acceptedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/355737
url http://hdl.handle.net/10261/355737
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/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/PID2022-137342OB-I00
info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/CEX 2021-001189-S
Applied and environmental microbiology
https://doi.org/10.1128/aem.02290-23
Sí
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.publisher.none.fl_str_mv American Society for Microbiology
publisher.none.fl_str_mv American Society for Microbiology
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
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spelling Ppx1 putative exopolyphosphatase is essential for polyphosphate accumulation in Lacticaseibacillus paracaseiCorrales, DanielaAlcántara, CristinaZúñiga, ManuelMonedero, VicenteLacticaseibacillus paracaseiExopolyphosphatasePolyphosphatePolyphosphate kinaselactic acid bacteriapolyphosphatesThe linear polymer polyphosphate (poly-P) is present across all three domains of life and serves diverse physiological functions. The enzyme polyphosphate kinase (Ppk) is responsible for poly-P synthesis, whereas poly-P degradation is carried out by the enzyme exopolyphosphatase (Ppx). In many Lactobacillaceae, the Ppk-encoding gene (ppk) is found clustered together with two genes encoding putative exopolyphosphatases (ppx1 and ppx2) each having different domain compositions, with the gene order ppx1-ppk-ppx2. However, the specific function of these ppx genes remains unexplored. An in-frame deletion of ppx1 in Lacticaseibacillus paracasei BL23 resulted in bacteria unable to accumulate poly-P, whereas the disruption of ppx2 did not affect poly-P synthesis. The expression of ppk was not altered in the Δppx1 strain, and poly-P synthesis in this strain was only restored by expressing ppx1 in trans. Moreover, no poly-P synthesis was observed when ppk was expressed from a plasmid in the Δppx1 strain. Purified Ppx2 exhibited in vitro exopolyphosphatase activity, whereas no in vitro enzymatic activity could be demonstrated for Ppx1. This observation corresponds with the absence in Ppx1 of conserved motifs essential for catalysis found in characterized exopolyphosphatases. Furthermore, assays with purified Ppk and Ppx1 evidenced that Ppx1 enhanced Ppk activity. These results demonstrate that Ppx1 is essential for poly-P synthesis in Lc. paracasei and have unveiled, for the first time, an unexpected role of Ppx1 exopolyphosphatase in poly-P synthesis.IMPORTANCEPoly-P is a pivotal molecular player in bacteria, participating in a diverse array of processes ranging from stress resilience to pathogenesis while also serving as a functional component in probiotic bacteria. The synthesis of poly-P is tightly regulated, but the underlying mechanisms remain incompletely elucidated. Our study sheds light on the distinctive role played by the two exopolyphosphatases (Ppx) found in the Lactobacillaceae bacterial group, of relevance in food and health. This particular group is noteworthy for possessing two Ppx enzymes, supposedly involved in poly-P degradation. Remarkably, our investigation uncovers an unprecedented function of Ppx1 in Lacticaseibacillus paracasei, where its absence leads to the total cessation of poly-P synthesis, paralleling the impact observed upon eliminating the poly-P forming enzyme, poly-P kinase. Unlike the anticipated role as a conventional exopolyphosphatase, Ppx1 demonstrates an unexpected function. Our results added a layer of complexity to our understanding of poly-P dynamics in bacteria.D. Corrales is grateful for her doctoral fellowship from the Ministerio de Ciencia, Tecnología e Innovación of Colombia (Convocatoria 906 de 2021, Doctorados en el Exterior). This research was partly funded by project PID2022-137342OB-100, MICIU/AEI/10.13039/501100011033/FEDER, UE and supported by the Severo Ochoa Accreditation CEX2021-001189-S from the MICIU, Spain.With funding from the Spanish government through the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX 2021-001189-S)Peer reviewedAmerican Society for MicrobiologyMinisterio de Ciencia, Tecnología e Innovación (Colombia)Agencia Estatal de Investigación (España)European CommissionConsejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]202420242024info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501Postprintinfo:eu-repo/semantics/acceptedVersionhttp://hdl.handle.net/10261/355737reponame: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/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/PID2022-137342OB-I00info:eu-repo/grantAgreement/AEI/Plan Estatal de Investigación Científica y Técnica y de Innovación 2021-2023/CEX 2021-001189-SApplied and environmental microbiologyhttps://doi.org/10.1128/aem.02290-23Síinfo:eu-repo/semantics/openAccessoai:digital.csic.es:10261/3557372026-05-22T06:33:51Z
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