Thermodynamic analysis of Kex2 activity: The acylation and deacylation steps are potassium- and substrate-dependent

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dc.citation.volumev. 235
dc.contributor.authorAntunes, Alyne Alexandrino
dc.contributor.authorJesus, Larissa de Oliveira Passos
dc.contributor.authorManfredi, Marcella Araujo
dc.contributor.authorSouza, Aline Aparecida de
dc.contributor.authorMachado, Mauricio Ferreira Marcondes
dc.contributor.authorSilva, Pamela Moraes e
dc.contributor.authorIcimoto, Marcelo Yudi [UNIFESP]
dc.contributor.authorJuliano, Maria Aparecida [UNIFESP]
dc.contributor.authorJuliano, Luiz [UNIFESP]
dc.contributor.authorJudice, Wagner Alves de Souza
dc.coverageAmsterdam
dc.date.accessioned2020-07-20T16:31:11Z
dc.date.available2020-07-20T16:31:11Z
dc.date.issued2018
dc.description.abstractKex2 is the prototype of a large family of eukaryotic subtilisin-related proprotein-processing proteases that cleave at sites containing pairs of basic residues. Here, we studied the effects of KCl on the individual rate constants of association, dissociation, acylation and deacylation and determined the thermodynamic parameters at each step of the Kex2 reaction. Potassium bound Kex2 with K-D = 20.3 mM. The order in which potassium entered the reaction system modified the effect of activation or inhibition, which depended on the size of the substrate. A possible allosteric potassium binding site at the S-6 subsite was involved in activation, and a distant site located between the catalytic domain and the P-domain was involved in inhibition. Potassium decreased the energetic barriers of almost all steps of catalysis. The acylation of Ac-PMYKR-AMC in the absence of potassium was the rate-limiting step. Therefore, for substrates containing a P-1-Arg, the deacylation step is not necessarily the rate-limiting event, and other residues at the P' positions may participate in controlling the acylation and deacylation steps. Thus, it is reasonable to conclude that potassium is involved in the processing of the alpha-mating factor that promotes Ca2+, mobilization by activating a high-affinity Ca2+-influx system to increase the cytosolic [Ca2+], resulting in the activation of channels that are essential for the survival of Saccharomyces cerevisine cells.en
dc.description.affiliationUniv Mogi Das Cruzes UMC, Ctr Interdisciplinar Invest Bioquim, Av Candido Xavier Almeida & Souza 200,Sala 1S-15, BR-08780911 Mogi Das Cruzes, SP, Brazil
dc.description.affiliationUniv Fed Sao Paulo, Escola Paulista Med, Dept Biofis, Rua Tres de Maio 100, BR-04044020 Sao Paulo, SP, Brazil
dc.description.affiliationUnifespUniv Fed Sao Paulo, Escola Paulista Med, Dept Biofis, Rua Tres de Maio 100, BR-04044020 Sao Paulo, SP, Brazil
dc.description.sourceWeb of Science
dc.description.sponsorshipFundacao de Amparo a Pesquisa do Estado de Sao Paulo - FAPESP
dc.description.sponsorshipIDFAPESP: 2014/02205-1
dc.description.sponsorshipIDFAPESP: 2016/25112-4
dc.description.sponsorshipIDFAPESP: 2015/11190-0
dc.format.extent29-39
dc.identifierhttp://dx.doi.org/10.1016/j.bpc.2017.11.007
dc.identifier.citationBiophysical Chemistry. Amsterdam, v. 235, p. 29-39, 2018.
dc.identifier.doi10.1016/j.bpc.2017.11.007
dc.identifier.issn0301-4622
dc.identifier.urihttps://repositorio.unifesp.br/handle/11600/55769
dc.identifier.wosWOS:000428008300004
dc.language.isoeng
dc.publisherElsevier Science Bv
dc.relation.ispartofBiophysical Chemistry
dc.rightsinfo:eu-repo/semantics/restrictedAccess
dc.subjectKex2en
dc.subjectSaccharomyces cerevisiaeen
dc.subjectThermodynamicsen
dc.subjectPotassium effectsen
dc.subjectAcylationen
dc.subjectDeacylationen
dc.titleThermodynamic analysis of Kex2 activity: The acylation and deacylation steps are potassium- and substrate-dependenten
dc.typeinfo:eu-repo/semantics/article
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