Mutations at the hydrophobic core affect Hal3 trimer stability, reducing its Ppz1 inhibitory capacity but not its PPCDC moonlighting function

Abstract S. cerevisiae Hal3 (ScHal3) is a moonlighting protein that, is in its monomeric state, regulates the Ser/Thr protein phosphatase Ppz1, but also joins ScCab3 (and in some instances the Hal3 paralog Vhs3) to form an unusual heterotrimeric phosphopantothenoylcysteine decarboxylase (PPCDC) enzy...

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Autores principales:	Carlos Santolaria, Diego Velázquez, Erick Strauss, Joaquín Ariño
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	Schally Phosphopantothenoylcysteine Decarboxylase Plasmid pWS12 Native AtHal3 Hall Variety Medicine R Science Q
Acceso en línea:	https://doaj.org/article/29a02033123b4902814ead3183ce1a04
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spelling	oai:doaj.org-article:29a02033123b4902814ead3183ce1a042021-12-02T15:07:52ZMutations at the hydrophobic core affect Hal3 trimer stability, reducing its Ppz1 inhibitory capacity but not its PPCDC moonlighting function10.1038/s41598-018-32979-x2045-2322https://doaj.org/article/29a02033123b4902814ead3183ce1a042018-10-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-32979-xhttps://doaj.org/toc/2045-2322Abstract S. cerevisiae Hal3 (ScHal3) is a moonlighting protein that, is in its monomeric state, regulates the Ser/Thr protein phosphatase Ppz1, but also joins ScCab3 (and in some instances the Hal3 paralog Vhs3) to form an unusual heterotrimeric phosphopantothenoylcysteine decarboxylase (PPCDC) enzyme. PPCDC is required for CoA biosynthesis and in most eukaryotes is a homotrimeric complex with three identical catalytic sites at the trimer interfaces. However, in S. cerevisiae the heterotrimeric arrangement results in a single functional catalytic center. Importantly, the specific structural determinants that direct Hal3’s oligomeric state and those required for Ppz1 inhibition remain largely unknown. We mutagenized residues in the predicted hydrophobic core of ScHal3 (L403–L405) and the plant Arabidopsis thaliana Hal3 (AtHal3, G115–L117) oligomers and characterized their properties as PPCDC components and, for ScHal3, also as Ppz1 inhibitor. We found that in AtHal3 these changes do not affect trimerization or PPCDC function. Similarly, mutation of ScHal3 L403 has no effect. In contrast, ScHal3 L405E fails to form homotrimers, but retains the capacity to bind Cab3—explaining its ability to rescue a hal3 vhs3 synthetically lethal mutation. Remarkably, the L405E mutation decreases Hal3’s ability to interact with and to inhibit Ppz1, confirming the importance of the oligomer/monomer equilibrium in Hal3’s Ppz1 regulating function.Carlos SantolariaDiego VelázquezErick StraussJoaquín AriñoNature PortfolioarticleSchallyPhosphopantothenoylcysteine DecarboxylasePlasmid pWS12Native AtHal3Hall VarietyMedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-12 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Schally Phosphopantothenoylcysteine Decarboxylase Plasmid pWS12 Native AtHal3 Hall Variety Medicine R Science Q
spellingShingle	Schally Phosphopantothenoylcysteine Decarboxylase Plasmid pWS12 Native AtHal3 Hall Variety Medicine R Science Q Carlos Santolaria Diego Velázquez Erick Strauss Joaquín Ariño Mutations at the hydrophobic core affect Hal3 trimer stability, reducing its Ppz1 inhibitory capacity but not its PPCDC moonlighting function
description	Abstract S. cerevisiae Hal3 (ScHal3) is a moonlighting protein that, is in its monomeric state, regulates the Ser/Thr protein phosphatase Ppz1, but also joins ScCab3 (and in some instances the Hal3 paralog Vhs3) to form an unusual heterotrimeric phosphopantothenoylcysteine decarboxylase (PPCDC) enzyme. PPCDC is required for CoA biosynthesis and in most eukaryotes is a homotrimeric complex with three identical catalytic sites at the trimer interfaces. However, in S. cerevisiae the heterotrimeric arrangement results in a single functional catalytic center. Importantly, the specific structural determinants that direct Hal3’s oligomeric state and those required for Ppz1 inhibition remain largely unknown. We mutagenized residues in the predicted hydrophobic core of ScHal3 (L403–L405) and the plant Arabidopsis thaliana Hal3 (AtHal3, G115–L117) oligomers and characterized their properties as PPCDC components and, for ScHal3, also as Ppz1 inhibitor. We found that in AtHal3 these changes do not affect trimerization or PPCDC function. Similarly, mutation of ScHal3 L403 has no effect. In contrast, ScHal3 L405E fails to form homotrimers, but retains the capacity to bind Cab3—explaining its ability to rescue a hal3 vhs3 synthetically lethal mutation. Remarkably, the L405E mutation decreases Hal3’s ability to interact with and to inhibit Ppz1, confirming the importance of the oligomer/monomer equilibrium in Hal3’s Ppz1 regulating function.
format	article
author	Carlos Santolaria Diego Velázquez Erick Strauss Joaquín Ariño
author_facet	Carlos Santolaria Diego Velázquez Erick Strauss Joaquín Ariño
author_sort	Carlos Santolaria
title	Mutations at the hydrophobic core affect Hal3 trimer stability, reducing its Ppz1 inhibitory capacity but not its PPCDC moonlighting function
title_short	Mutations at the hydrophobic core affect Hal3 trimer stability, reducing its Ppz1 inhibitory capacity but not its PPCDC moonlighting function
title_full	Mutations at the hydrophobic core affect Hal3 trimer stability, reducing its Ppz1 inhibitory capacity but not its PPCDC moonlighting function
title_fullStr	Mutations at the hydrophobic core affect Hal3 trimer stability, reducing its Ppz1 inhibitory capacity but not its PPCDC moonlighting function
title_full_unstemmed	Mutations at the hydrophobic core affect Hal3 trimer stability, reducing its Ppz1 inhibitory capacity but not its PPCDC moonlighting function
title_sort	mutations at the hydrophobic core affect hal3 trimer stability, reducing its ppz1 inhibitory capacity but not its ppcdc moonlighting function
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/29a02033123b4902814ead3183ce1a04
work_keys_str_mv	AT carlossantolaria mutationsatthehydrophobiccoreaffecthal3trimerstabilityreducingitsppz1inhibitorycapacitybutnotitsppcdcmoonlightingfunction AT diegovelazquez mutationsatthehydrophobiccoreaffecthal3trimerstabilityreducingitsppz1inhibitorycapacitybutnotitsppcdcmoonlightingfunction AT erickstrauss mutationsatthehydrophobiccoreaffecthal3trimerstabilityreducingitsppz1inhibitorycapacitybutnotitsppcdcmoonlightingfunction AT joaquinarino mutationsatthehydrophobiccoreaffecthal3trimerstabilityreducingitsppz1inhibitorycapacitybutnotitsppcdcmoonlightingfunction
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Mutations at the hydrophobic core affect Hal3 trimer stability, reducing its Ppz1 inhibitory capacity but not its PPCDC moonlighting function

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