Properties of Subunits of the Multicatalytic Proteinase Complex Revealed by the Use of Subunit-specific Antibodies

Overview

Journal Biochem J

Specialty Biochemistry

Date 1991 Aug 15

PMID 1883328

Citations 9

Authors

A J Rivett

S T Sweeney

Affiliations

Soon will be listed here.

Abstract

The multicatalytic proteinase (MCP) is a high-molecular-mass non-lysosomal proteinase that gives rise to a characteristic pattern of bands of molecular mass 22-34 kDa on SDS/PAGE gels. Isoelectric-focusing gels of the enzyme purified from rat liver show 16 bands with isoelectric points in the range of pH 5-8.5. Two-dimensional PAGE gels reveal that there are more than the previously reported 13 polypeptides associated with the MCP from rat liver and show a pattern of 15-20 major spots and several minor ones, similar to that of MCP isolated from some other sources. Possible relationships between the different polypeptides were investigated by immunoblot analysis of electrophoretically purified proteinase subunits with affinity-purified subunit-specific antibodies as well as antibodies raised against individual denatured subunits of the complex. The results demonstrate that many of the major polypeptide components of the MCP complex are antigenically distinct. Moreover comparison of immunoreactive material in crude cell extracts with that in purified MCP preparations has shown that the polypeptides are not derived from a smaller number of higher-molecular-mass subunits. Also, individual subunits have the same apparent molecular mass in a variety of rat tissues, suggesting close similarity between MCPs of different tissues. The highest concentrations of MCP subunits occur in liver and kidney. Gel-filtration analysis of crude extracts has demonstrated that MCP polypeptides are also associated with a higher-molecular-mass complex, which may be the 26 S proteinase that has been implicated in the degradation of ubiquitin-protein conjugates.

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References

McGuire M, Croall D, DeMartino G . ATP-stimulated proteolysis in soluble extracts of BHK 21/C13 cells. Evidence for multiple pathways and a role for an enzyme related to the high-molecular-weight protease, macropain. Arch Biochem Biophys. 1988; 262(1):273-85. DOI: 10.1016/0003-9861(88)90189-0. View

Ray K, Harris H . Lens neutral endopeptidase occurs in other bovine and human tissues. Biochem J. 1987; 248(3):643-8. PMC: 1148597. DOI: 10.1042/bj2480643. View

Rivett A . Purification of a liver alkaline protease which degrades oxidatively modified glutamine synthetase. Characterization as a high molecular weight cysteine proteinase. J Biol Chem. 1985; 260(23):12600-6. View

Martins de Sa C, Grossi de Sa M, Akhayat O, Broders F, Scherrer K, Horsch A . Prosomes. Ubiquity and inter-species structural variation. J Mol Biol. 1986; 187(4):479-93. DOI: 10.1016/0022-2836(86)90328-1. View

Ray K, Harris H . Comparative studies on lens neutral endopeptidase and pituitary neutral proteinase: two closely similar enzymes. FEBS Lett. 1986; 194(1):91-5. DOI: 10.1016/0014-5793(86)80057-6. View

Tanaka K, Ii K, Ichihara A, Waxman L, GOLDBERG A . A high molecular weight protease in the cytosol of rat liver. I. Purification, enzymological properties, and tissue distribution. J Biol Chem. 1986; 261(32):15197-203. View

Tanaka K, Yoshimura T, Ichihara A, Ikai A, Nishigai M, Morimoto Y . Molecular organization of a high molecular weight multi-protease complex from rat liver. J Mol Biol. 1988; 203(4):985-96. DOI: 10.1016/0022-2836(88)90123-4. View

Arrigo A, Tanaka K, GOLDBERG A, Welch W . Identity of the 19S 'prosome' particle with the large multifunctional protease complex of mammalian cells (the proteasome). Nature. 1988; 331(6152):192-4. DOI: 10.1038/331192a0. View

Tomek W, Adam G, Schmid H . Prosomes, small cytoplasmic RNP particles, contain glycoproteins. FEBS Lett. 1988; 239(1):155-8. DOI: 10.1016/0014-5793(88)80564-7. View

10.

Grossi de Sa M, Martins de Sa C, Harper F, Coux O, Akhayat O, Pal J . Cytolocalization of prosomes as a function of differentiation. J Cell Sci. 1988; 89 ( Pt 2):151-65. DOI: 10.1242/jcs.89.2.151. View

11.

Hough R, Pratt G, Rechsteiner M . Purification of two high molecular weight proteases from rabbit reticulocyte lysate. J Biol Chem. 1987; 262(17):8303-13. View

12.

Haass C, Multhaup G, Beyreuther K, Kloetzel P . The Drosophila PROS-28.1 gene is a member of the proteasome gene family. Gene. 1990; 90(2):235-41. DOI: 10.1016/0378-1119(90)90185-t. View

13.

Driscoll J, GOLDBERG A . The proteasome (multicatalytic protease) is a component of the 1500-kDa proteolytic complex which degrades ubiquitin-conjugated proteins. J Biol Chem. 1990; 265(9):4789-92. View

14.

Kumatori A, Tanaka K, Tamura T, Fujiwara T, Ichihara A, Tokunaga F . cDNA cloning and sequencing of component C9 of proteasomes from rat hepatoma cells. FEBS Lett. 1990; 264(2):279-82. DOI: 10.1016/0014-5793(90)80267-m. View

15.

Lilley K, Davison M, Rivett A . N-terminal sequence similarities between components of the multicatalytic proteinase complex. FEBS Lett. 1990; 262(2):327-9. DOI: 10.1016/0014-5793(90)80220-d. View

16.

Lee L, Moomaw C, Orth K, McGuire M, DeMartino G, Slaughter C . Relationships among the subunits of the high molecular weight proteinase, macropain (proteasome). Biochim Biophys Acta. 1990; 1037(2):178-85. DOI: 10.1016/0167-4838(90)90165-c. View

17.

Duncan R, Hershey J . Evaluation of isoelectric focusing running conditions during two-dimensional isoelectric focusing/sodium dodecyl sulfate-polyacrylamide gel electrophoresis: variation of gel patterns with changing conditions and optimized isoelectric focusing.... Anal Biochem. 1984; 138(1):144-55. DOI: 10.1016/0003-2697(84)90783-8. View

18.

Bos E, van der Doelen A, van Rooy N, SCHUURS A . 3,3',5,5' - Tetramethylbenzidine as an Ames test negative chromogen for horse-radish peroxidase in enzyme-immunoassay. J Immunoassay. 1981; 2(3-4):187-204. DOI: 10.1080/15321818108056977. View

19.

Towbin H, Staehelin T, Gordon J . Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979; 76(9):4350-4. PMC: 411572. DOI: 10.1073/pnas.76.9.4350. View

20.

Ofarrell P . High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975; 250(10):4007-21. PMC: 2874754. View