The presence of ATP + ubiquitin-dependent proteinase and multicatalytic proteinase complex in bovine brain.
The presence of two distinct high-molecular-weight proteases with similar pH optima in the weakly alkaline region was shown in cytosol of the bovine brain cortex. They were separated by ammonium sulfate fractionation and each was further purified by DEAE-Sephacel, Sephacryl S-300, DEAE-Cibacron Blue 3GA-agarose, heparin-agarose, and Sepharose 6B chromatography. The larger enzyme (Mr 1,400 kDa), which precipitates at 0-38% ammonium sulfate saturation, seems to be active in ATP + ubiquitin (Ub)-dependent proteolysis; it has low basal caseinolytic activity that is stimulated 3-fold by ATP, and when Ub is present ATP causes a 4.5-fold stimulation. A second proteinase was also found to be present (Mr 700 kDa) that precipitates at 38-80% ammonium sulfate saturation, is composed of multiple subunits ranging in Mr from 18 to 30 kDa, and degrades both protein and peptide substrates, demonstrating trypsin-, chymotrypsin- and cucumisin-like activities. Catalytic, biochemical, and immunological characteristics of this proteinase indicate that it is a multicatalytic proteinase complex (MPC), whose enzyme activity, in contrast to that of MPC from bovine pituitaries (1-3), is stimulated 1.7-fold by addition of ATP in the absence of ubiquitin at the early steps of purification; this property is lost during the course of further purification. Both proteinases are present in the nerve cells, since the primary chicken embryonic telencephalon neuronal cell culture extracts contain both ATP + Ub-dependent proteinase and MPC activities.
Azaryan
A
A
Institute of Biochemistry, Arm. SSR Acad. Sci., Yerevan, USSR.
Banay-Schwartz
M
M
Lajtha
A
A
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D002540:Cerebral Cortex / Q000201:enzymology*; D003546:Cysteine Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D009097:Multienzyme Complexes / Q000302:isolation & purification* / Q000378:metabolism; D010447:Peptide Hydrolases / Q000302:isolation & purification* / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex; D013379:Substrate Specificity; D014452:Ubiquitins / Q000494:pharmacology*
D009097:Multienzyme Complexes; D014452:Ubiquitins; D000255:Adenosine Triphosphate; D010447:Peptide Hydrolases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex