title: Identification and purification of a 90-kDa membrane-bound endogenous inhibitor of multicatalytic proteinase from human erythrocytes.
abstract: We have identified and purified an endogenous inhibitor of multicatalytic proteinase (MCP) from human erythrocyte membranes. The inhibitor showed a molecular mass of 90 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The inhibitor protein was purified from the erythrocyte membranes using Heparinagarose and hydroxylapatite chromatography and the size exclusion on a Biogel A 1.5 m column in the presence of high salt. The 90-kDa protein inhibited all three peptidase activities of MCP; trypsin-like, chymotrypsin-like and peptidyl glutamyl peptide hydrolyzing (PGPH). However, it failed to cause any significant inhibition of caseinolytic activity of MCP, suggesting that the regulation of proteinase and peptidase activities is distinct. The inhibition of the chymotrypsin-like activity was noncompetitive. The results suggest that the 90-kDa inhibitor protein may be an important regulator of membrane-bound MCP.
authors.affiliation: Department of Pharmacy, University of Sydney, NSW, Australia.
authors.forename: M T
authors.initials: MT
authors.lastname: Khan
authors.forename: K
authors.initials: K
authors.lastname: Wang
authors.forename: B D
authors.initials: BD
authors.lastname: Roufogalis
mesh_terms: D000595:Amino Acid Sequence; D002845:Chromatography; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D003546:Cysteine Endopeptidases / Q000097:blood* / Q000302:isolation & purification; D017886:Durapatite; D004591:Electrophoresis, Polyacrylamide Gel; D004910:Erythrocyte Membrane / Q000378:metabolism*; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000097:blood* / Q000302:isolation & purification / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009097:Multienzyme Complexes / Q000097:blood* / Q000302:isolation & purification; D011480:Protease Inhibitors / Q000097:blood* / Q000302:isolation & purification / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex; D013379:Substrate Specificity
chemical_list: D008565:Membrane Proteins; D009097:Multienzyme Complexes; D011480:Protease Inhibitors; D017886:Durapatite; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex