Isolation and characterization of cathepsin Z, a lysosomal cysteine proteinase

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dc.contributor.authorDeCourcy, Kristi R.en
dc.contributor.committeecochairMason, Robert W.en
dc.contributor.committeecochairStorrie, Brianen
dc.contributor.committeememberBender, Patrick K.en
dc.contributor.committeememberNewton, William E.en
dc.contributor.committeememberRutherford, Charles L.en
dc.contributor.departmentBiochemistry and Anaerobic Microbiologyen
dc.date.accessioned2014-03-14T21:18:22Zen
dc.date.adate2007-08-27en
dc.date.available2014-03-14T21:18:22Zen
dc.date.issued1995-05-05en
dc.date.rdate2007-08-27en
dc.date.sdate2007-08-27en
dc.description.abstractCathepsin Z is a cysteine proteinase found in lysosomes of human cells. It was detected in human cultured cell lines using the peptidyl diazomethane inhibitor Fmoc-Leu-Leu- [¹²⁵]Tyr-CHN₂. The labeling of cathepsin Z by the inhibitor was both time- and concentration-dependent, and the proteinase was found in all human cell lines examined. The characteristics of cathepsin Z were examined in U-937 cells, a human monocytic line. The labeling of cathepsin Z was blocked by pre-incubation of the cells either in non-iodinated inhibitor or in the epoxysuccinyl peptide inhibitor E-64d, a specific inhibitor of cysteine proteinases. Cathepsin Z was not immunoprecipitated by antisera specific for cathepsins B, L, or S. Cathepsin Z has been estimated to be at millimolar concentrations in lysosomes, suggesting that it is a major lysosomal proteinase. The molecular weight of cathepsin Z was calculated to be 22.4 kDa by SDS-PAGE and 45— 47 kDa by native PAGE and gel exclusion chromatography, indicating that it is dimeric. Cathepsin Z is susceptible to digestion by endoglycosidase H, and oligosaccharides comprise 3.1 kDa of the reduced molecular weight. The expression of cathepsin Z was not affected by differentiation of U-937 cells with phorbol ester, unlike the expression of cathepsins B and S. Undifferentiated U-937 cells express low levels of cathepsins B and S; after differentiation, expression of cathepsins B and S is greatly increased. Cathepsin Z was purified from U-937 cells by anion exchange chromatography (Mono Q), affinity chromatography (concanavalin A), and preparatory electrophoresis. N-terminal sequence analysis of both the purified protein and fragments of the protein from a V8 digest indicates that cathepsin Z is a member of the papain superfamily of cysteine proteinases.en
dc.description.degreePh. D.en
dc.format.extentxiv, 186 leavesen
dc.format.mediumBTDen
dc.format.mimetypeapplication/pdfen
dc.identifier.otheretd-08272007-163757en
dc.identifier.sourceurlhttp://scholar.lib.vt.edu/theses/available/etd-08272007-163757/en
dc.identifier.urihttp://hdl.handle.net/10919/39271en
dc.language.isoenen
dc.publisherVirginia Techen
dc.relation.haspartLD5655.V856_1995.D436.pdfen
dc.relation.isformatofOCLC# 32883586en
dc.rightsIn Copyrighten
dc.rights.urihttp://rightsstatements.org/vocab/InC/1.0/en
dc.subjecthuman cell linesen
dc.subject.lccLD5655.V856 1995.D436en
dc.titleIsolation and characterization of cathepsin Z, a lysosomal cysteine proteinaseen
dc.typeDissertationen
dc.type.dcmitypeTexten
thesis.degree.disciplineBiochemistry and Anaerobic Microbiologyen
thesis.degree.grantorVirginia Polytechnic Institute and State Universityen
thesis.degree.leveldoctoralen
thesis.degree.namePh. D.en

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