Recombinant expression, purification, and functional analysis of two novel cystatins from sugarcane (Saccharum officinarum)

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dc.contributor.authorGianotti, A.
dc.contributor.authorRios, W. M.
dc.contributor.authorSoares-Costa, A.
dc.contributor.authorNogaroto, V
dc.contributor.authorCarmona, A. K.
dc.contributor.authorOliva, MLV
dc.contributor.authorAndrade, S. S.
dc.contributor.authorHenrique-Silva, F.
dc.contributor.institutionUniversidade Federal de São Carlos (UFSCar)
dc.contributor.institutionUniversidade Federal de São Paulo (UNIFESP)
dc.date.accessioned2016-01-24T12:41:14Z
dc.date.available2016-01-24T12:41:14Z
dc.date.issued2006-06-01
dc.description.abstractPhytocystatins are cysteine proteinase inhibitors from plants implicated in the endogenous regulation of protein turnover, programmed cell death, and in defense mechanisms against pathogens. To date, only few cystatin genes have been characterized in most plant species. We have previously characterized the protein Canecystatin, the first cystatin described in sugarcane. in an attempt to study novel Canecystatins, we identified two ORFs encoding cystatins (referred as CaneCPI-2 and CaneCPI-3) using the data from the Sugarcane EST genome project. These ORFs were then subcloned and expressed in Escherichia coli using pET28 expression vector. High amounts (similar to 20 mg/L) of pure recombinant proteins were obtained by affinity chromatography in a single step of purification. Polyclonal antibodies against the recombinant Canecystatins were raised, allowing the immunodetection of the endogenous proteins in the plant tissues. Moreover, the proteins were able to inhibit papain in a fluorometric assay with K-i values of 0.2 and 0.25 mu M for CaneCPI-2 and CaneCPI-3, respectively. These findings contribute to a better understanding of the activity of sugarcane cystatins and encourage future activity and structural studies of these proteins. (c) 2005 Elsevier Inc. All rights reserved.en
dc.description.affiliationUniv Fed Sao Carlos, DGE, Lab Mol Biol, BR-13565905 Sao Carlos, SP, Brazil
dc.description.affiliationUniversidade Federal de São Paulo, Escola Paulista Med, Dept Biofis, BR-04044020 São Paulo, Brazil
dc.description.affiliationUniversidade Federal de São Paulo, Dept Bioquim, Escola Paulista Med, BR-04044020 São Paulo, Brazil
dc.description.affiliationUnifespUniversidade Federal de São Paulo, Escola Paulista Med, Dept Biofis, BR-04044020 São Paulo, Brazil
dc.description.affiliationUnifespUniversidade Federal de São Paulo, Dept Bioquim, Escola Paulista Med, BR-04044020 São Paulo, Brazil
dc.description.sourceWeb of Science
dc.format.extent483-489
dc.identifierhttp://dx.doi.org/10.1016/j.pep.2005.10.026
dc.identifier.citationProtein Expression and Purification. San Diego: Academic Press Inc Elsevier Science, v. 47, n. 2, p. 483-489, 2006.
dc.identifier.doi10.1016/j.pep.2005.10.026
dc.identifier.issn1046-5928
dc.identifier.urihttp://repositorio.unifesp.br/handle/11600/28956
dc.identifier.wosWOS:000238277000020
dc.language.isoeng
dc.publisherElsevier B.V.
dc.relation.ispartofProtein Expression and Purification
dc.rightsinfo:eu-repo/semantics/restrictedAccess
dc.rights.licensehttp://www.elsevier.com/about/open-access/open-access-policies/article-posting-policy
dc.subjectcysteine proteinase inhibitoren
dc.subjectcystatinen
dc.subjectphytocystatinen
dc.subjectcanecystatinen
dc.subjectsugarcaneen
dc.subjectSaccharum officinarumen
dc.titleRecombinant expression, purification, and functional analysis of two novel cystatins from sugarcane (Saccharum officinarum)en
dc.typeinfo:eu-repo/semantics/article
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