Antifungal Activity of the <named-content content-type="genus-species">Enterococcus faecalis</named-content> Peptide EntV Requires Protease Cleavage and Disulfide Bond Formation

ABSTRACT Enterococcus faecalis, a Gram-positive bacterium, and Candida albicans, a polymorphic fungus, are common constituents of the microbiome as well as increasingly problematic causes of infections. Interestingly, we previously showed that these two species antagonize each other’s virulence and...

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Autores principales:	Armand O. Brown, Carrie E. Graham, Melissa R. Cruz, Kavindra V. Singh, Barbara E. Murray, Michael C. Lorenz, Danielle A. Garsin
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	Candida Enterococcus antifungals bacteriocins Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/3770f5e545754a4a8fdfd990b4b0f751
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spelling	oai:doaj.org-article:3770f5e545754a4a8fdfd990b4b0f7512021-11-15T16:22:10ZAntifungal Activity of the <named-content content-type="genus-species">Enterococcus faecalis</named-content> Peptide EntV Requires Protease Cleavage and Disulfide Bond Formation10.1128/mBio.01334-192150-7511https://doaj.org/article/3770f5e545754a4a8fdfd990b4b0f7512019-08-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01334-19https://doaj.org/toc/2150-7511ABSTRACT Enterococcus faecalis, a Gram-positive bacterium, and Candida albicans, a polymorphic fungus, are common constituents of the microbiome as well as increasingly problematic causes of infections. Interestingly, we previously showed that these two species antagonize each other’s virulence and that E. faecalis inhibition of C. albicans was specifically mediated by EntV. EntV is a bacteriocin encoded by the entV (ef1097) locus that reduces C. albicans virulence and biofilm formation by inhibiting hyphal morphogenesis. In this report, we studied the posttranslational modifications necessary for EntV antifungal activity. First, we show that the E. faecalis secreted enzyme gelatinase (GelE) is responsible for cleaving EntV into its 68-amino-acid, active form and that this process does not require the serine protease SprE. Furthermore, we demonstrate that a disulfide bond that forms within EntV is necessary for antifungal activity. Abrogating this bond by chemical treatment or genetic modification rendered EntV inactive against C. albicans. Moreover, we identified the likely catalyst of this disulfide bond, a previously uncharacterized thioredoxin within the E. faecalis genome called DsbA. Loss of DsbA, or disruption of its redox-active cysteines, resulted in loss of EntV antifungal activity. Finally, we show that disulfide bond formation is not a prerequisite for cleavage; EntV cleavage proceeded normally in the absence of DsbA. In conclusion, we present a model in which following secretion, EntV undergoes disulfide bond formation by DsbA and cleavage by GelE in order to generate a peptide capable of inhibiting C. albicans. IMPORTANCE Enterococcus faecalis and Candida albicans are among the most important and problematic pathobionts, organisms that normally are harmless commensals but can cause dangerous infections in immunocompromised hosts. In fact, both organisms are listed by the Centers for Disease Control and Prevention as serious global public health threats stemming from the increased prevalence of antimicrobial resistance. The rise in antifungal resistance is of particular concern considering the small arsenal of currently available therapeutics. EntV is a peptide with antifungal properties, and it, or a similar compound, could be developed into a therapeutic alternative, either alone or in combination with existing agents. However, to do so requires understanding what properties of EntV are necessary for its antifungal activity. In this work, we studied the posttranslational processing of EntV and what modifications are necessary for inhibition of C. albicans in order to fill this gap in knowledge.Armand O. BrownCarrie E. GrahamMelissa R. CruzKavindra V. SinghBarbara E. MurrayMichael C. LorenzDanielle A. GarsinAmerican Society for MicrobiologyarticleCandidaEnterococcusantifungalsbacteriocinsMicrobiologyQR1-502ENmBio, Vol 10, Iss 4 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	Candida Enterococcus antifungals bacteriocins Microbiology QR1-502
spellingShingle	Candida Enterococcus antifungals bacteriocins Microbiology QR1-502 Armand O. Brown Carrie E. Graham Melissa R. Cruz Kavindra V. Singh Barbara E. Murray Michael C. Lorenz Danielle A. Garsin Antifungal Activity of the <named-content content-type="genus-species">Enterococcus faecalis</named-content> Peptide EntV Requires Protease Cleavage and Disulfide Bond Formation
description	ABSTRACT Enterococcus faecalis, a Gram-positive bacterium, and Candida albicans, a polymorphic fungus, are common constituents of the microbiome as well as increasingly problematic causes of infections. Interestingly, we previously showed that these two species antagonize each other’s virulence and that E. faecalis inhibition of C. albicans was specifically mediated by EntV. EntV is a bacteriocin encoded by the entV (ef1097) locus that reduces C. albicans virulence and biofilm formation by inhibiting hyphal morphogenesis. In this report, we studied the posttranslational modifications necessary for EntV antifungal activity. First, we show that the E. faecalis secreted enzyme gelatinase (GelE) is responsible for cleaving EntV into its 68-amino-acid, active form and that this process does not require the serine protease SprE. Furthermore, we demonstrate that a disulfide bond that forms within EntV is necessary for antifungal activity. Abrogating this bond by chemical treatment or genetic modification rendered EntV inactive against C. albicans. Moreover, we identified the likely catalyst of this disulfide bond, a previously uncharacterized thioredoxin within the E. faecalis genome called DsbA. Loss of DsbA, or disruption of its redox-active cysteines, resulted in loss of EntV antifungal activity. Finally, we show that disulfide bond formation is not a prerequisite for cleavage; EntV cleavage proceeded normally in the absence of DsbA. In conclusion, we present a model in which following secretion, EntV undergoes disulfide bond formation by DsbA and cleavage by GelE in order to generate a peptide capable of inhibiting C. albicans. IMPORTANCE Enterococcus faecalis and Candida albicans are among the most important and problematic pathobionts, organisms that normally are harmless commensals but can cause dangerous infections in immunocompromised hosts. In fact, both organisms are listed by the Centers for Disease Control and Prevention as serious global public health threats stemming from the increased prevalence of antimicrobial resistance. The rise in antifungal resistance is of particular concern considering the small arsenal of currently available therapeutics. EntV is a peptide with antifungal properties, and it, or a similar compound, could be developed into a therapeutic alternative, either alone or in combination with existing agents. However, to do so requires understanding what properties of EntV are necessary for its antifungal activity. In this work, we studied the posttranslational processing of EntV and what modifications are necessary for inhibition of C. albicans in order to fill this gap in knowledge.
format	article
author	Armand O. Brown Carrie E. Graham Melissa R. Cruz Kavindra V. Singh Barbara E. Murray Michael C. Lorenz Danielle A. Garsin
author_facet	Armand O. Brown Carrie E. Graham Melissa R. Cruz Kavindra V. Singh Barbara E. Murray Michael C. Lorenz Danielle A. Garsin
author_sort	Armand O. Brown
title	Antifungal Activity of the <named-content content-type="genus-species">Enterococcus faecalis</named-content> Peptide EntV Requires Protease Cleavage and Disulfide Bond Formation
title_short	Antifungal Activity of the <named-content content-type="genus-species">Enterococcus faecalis</named-content> Peptide EntV Requires Protease Cleavage and Disulfide Bond Formation
title_full	Antifungal Activity of the <named-content content-type="genus-species">Enterococcus faecalis</named-content> Peptide EntV Requires Protease Cleavage and Disulfide Bond Formation
title_fullStr	Antifungal Activity of the <named-content content-type="genus-species">Enterococcus faecalis</named-content> Peptide EntV Requires Protease Cleavage and Disulfide Bond Formation
title_full_unstemmed	Antifungal Activity of the <named-content content-type="genus-species">Enterococcus faecalis</named-content> Peptide EntV Requires Protease Cleavage and Disulfide Bond Formation
title_sort	antifungal activity of the <named-content content-type="genus-species">enterococcus faecalis</named-content> peptide entv requires protease cleavage and disulfide bond formation
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/3770f5e545754a4a8fdfd990b4b0f751
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Antifungal Activity of the <named-content content-type="genus-species">Enterococcus faecalis</named-content> Peptide EntV Requires Protease Cleavage and Disulfide Bond Formation

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