Biochemical and Functional Characterization of a Metalloprotease from the Thermophilic Fungus Thermoascus aurantiacus

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dc.contributor.authorMerheb-Dini, Carolina [UNESP]
dc.contributor.authorCabral, Hamilton [UNESP]
dc.contributor.authorLeite, Rodrigo S. R. [UNESP]
dc.contributor.authorZanphorlin, Leticia M. [UNESP]
dc.contributor.authorOkamoto, Debora N. [UNESP]
dc.contributor.authorBonilla-Rodriguez, Gustavo Orlando [UNESP]
dc.contributor.authorJuliano, Luiz
dc.contributor.authorArantes, Eliane C.
dc.contributor.authorGomes, Eleni [UNESP]
dc.contributor.authorda Silva, Roberto [UNESP]
dc.contributor.institutionUniversidade Estadual Paulista (Unesp)
dc.contributor.institutionUniversidade de São Paulo (USP)
dc.contributor.institutionUniversidade Federal de São Paulo (UNIFESP)
dc.date.accessioned2014-05-20T14:01:08Z
dc.date.available2014-05-20T14:01:08Z
dc.date.issued2009-10-14
dc.description.abstractProtease production was carried out in solid state fermentation. The enzyme was purified through precipitation with ethanol at 72% followed by chromatographies in columns of Sephadex G75 and Sephacryl S100. It was purified 80-fold and exhibited recovery of total activity of 0.4%. SDS-PAGE analysis indicated an estimated molecular mass of 24.5 kDa and the N-terminal sequence of the first 22 residues was APYSGYQCSMQLCLTCALMNCA. Purified protease was only inhibited by EDTA (96.7%) and stimulated by Fe(2+) revealing to be a metalloprotease activated by iron. Optimum pH was 5.5, optimum temperature was 75 degrees C, and it was thermostable at 65 degrees C for 1 h maintaining more than 70% of original activity. Through enzyme kinetic studies, protease better hydrolyzed casein than azocasein. The screening of fluorescence resonance energy transfer (FRET) peptide series derived from Abz-KLXSSKQ-EDDnp revealed that the enzyme exhibited preference for Arg in P(1) (k(cat)/K(m) = 30.1 mM(-1) s(-1)).en
dc.description.affiliationUniv Estadual Paulista, Lab Bioquim & Microbiol Aplicada, Inst Biociencias Letras & Ciencias Exatas, BR-15054000 São Paulo, Brazil
dc.description.affiliationUniv São Paulo, Fac Ciencias Farmaceut Ribeirao Preto, BR-14040903 São Paulo, Brazil
dc.description.affiliationUniv Fed São Paulo, Dept Biofis, Escola Paulista Med, BR-04044020 São Paulo, Brazil
dc.description.affiliationUnespUniv Estadual Paulista, Lab Bioquim & Microbiol Aplicada, Inst Biociencias Letras & Ciencias Exatas, BR-15054000 São Paulo, Brazil
dc.description.sponsorshipFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
dc.description.sponsorshipConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)
dc.description.sponsorshipIdCNPq: 151868/2006-9
dc.format.extent9210-9217
dc.identifierhttp://dx.doi.org/10.1021/jf9017977
dc.identifier.citationJournal of Agricultural and Food Chemistry. Washington: Amer Chemical Soc, v. 57, n. 19, p. 9210-9217, 2009.
dc.identifier.doi10.1021/jf9017977
dc.identifier.issn0021-8561
dc.identifier.lattes6955258588672130
dc.identifier.lattes7091241742851920
dc.identifier.lattes9424175688206545
dc.identifier.urihttp://hdl.handle.net/11449/21609
dc.identifier.wosWOS:000270461500067
dc.language.isoeng
dc.publisherAmer Chemical Soc
dc.relation.ispartofJournal of Agricultural and Food Chemistry
dc.relation.ispartofjcr3.412
dc.relation.ispartofsjr1,269
dc.rights.accessRightsAcesso restrito
dc.sourceWeb of Science
dc.subjectThermoascus aurantiacusen
dc.subjectSolid state fermentation (SSF)en
dc.subjectmetalloproteaseen
dc.subjectThermostabilityen
dc.subjectfluorescent peptidesen
dc.subjectN-terminal sequenceen
dc.titleBiochemical and Functional Characterization of a Metalloprotease from the Thermophilic Fungus Thermoascus aurantiacusen
dc.typeArtigo
dcterms.licensehttp://pubs.acs.org/page/policy/nih/index.html
dcterms.rightsHolderAmer Chemical Soc
unesp.author.lattes6955258588672130
unesp.author.lattes7091241742851920
unesp.author.lattes9424175688206545
unesp.campusUniversidade Estadual Paulista (Unesp), Instituto de Biociências Letras e Ciências Exatas, São José do Rio Pretopt

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Artigos - Biologia - IBILCE