Isolation and characterization of a novel endogenous inhibitor of the proteasome
| Autor: | Joseph D. Etlinger, Xiaochong Li, Maozhi Gu |
|---|---|
| Rok vydání: | 1991 |
| Předmět: |
Erythrocytes
medicine.medical_treatment Proteolysis Molecular Sequence Data Biochemistry Catalysis Mice Ubiquitin Western blot medicine Animals Humans Protease Inhibitors Amino Acid Sequence Peptide sequence Mice Inbred BALB C Protease Molecular mass biology medicine.diagnostic_test Chemistry Antibodies Monoclonal Molecular biology Molecular Weight Proteasome Enzyme inhibitor biology.protein Rabbits |
| Zdroj: | Biochemistry. 30(40) |
| ISSN: | 0006-2960 7588-7592 |
| Popis: | A novel endogenous inhibitor of the proteasome (high molecular weight multicatalytic protease) has been isolated and characterized from human erythrocytes. After purification by ion-exchange and sizing chromatography, the inhibitor displayed a native molecular mass of approximately 200 kDa and contained a single subunit of 50 kDa with an isoelectric point of 6.9. Although the inhibitor noncompetitively blocks proteolysis of [methyl-14C]-alpha-casein (Ki = 7.1 x 10(-8) M) and inhibits hydrolysis of Suc-Leu-Leu-Val-Tyr-AMC, it did not affect hydrolysis of other peptide substrates, such as MeOSuc-Phe-Leu-Phe-MNA and Z-Ala-Arg-Arg-MNA. To further characterize the 50-kDa inhibitor, a monoclonal antibody (MI-8) was generated that showed specific binding upon Western blot analysis of both native PAGE and SDS-PAGE. Immunoprecipitation with MI-8 specifically removed inhibitor activity against the proteasome. The 50-kDa inhibitor is distinct from a previously described 40-kDa inhibitor of the proteasome (Murakami, K., & Etlinger, J.D. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 7588-7592) on the basis of lack of cross-reactivity with MI-8 and dissimilar peptide digest patterns. It is suggested that these endogenous inhibitors may have a role in ATP/ubiquitin-dependent proteolysis and/or other cellular functions involving this protease. |
| Databáze: | OpenAIRE |
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