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Schlagwörter:
Amino Acid Sequence; Cations, Divalent/ch [Chemistry]; Cations, Monovalent/ch [Chemistry]; Coumarins/pd [Pharmacology]; *Cysteine Endopeptidases/ch [Chemistry]; Enzyme Activation/de [Drug Effects]; Hydrogen-Ion Concentration; Isoelectric Point; Molecular Sequence Data; Molecular Weight; *Multienzyme Complexes/ch [Chemistry]; Peptides/ch [Chemistry]; Peptides/me [Metabolism]; Serpins/pd [Pharmacology]; Support, Non-U.S. Gov't; Temperature; Thermoplasma/ch [Chemistry]; *Thermoplasma/en [Enzymology]
Zusammenfassung:
We have purified proteasomes to apparent homogeneity from the archaebacterium Thermoplasma acidophilum. This proteinase has a molecular mass of about 650 kDa and an isoelectric point of 5.6. The proteasome hydrolyses peptide substrates containing an aromatic residue adjacent to the reporter group, as well as and 2 over black square]; [1 and 2 over black square]4C]methylated casein optimally at pH 8.5 and 90 degrees C. The enzyme activity is enhanced severalfold by Mg2+ and Ca2+ at 25-500 mM. This increase in activity results primarily from a change in Km. The serine-proteinase inhibitors diisopropylfluorophosphate and 3,4-dichloroisocoumarin irreversibly inhibit the enzyme, obviously by modification of both the alpha and beta subunits in the proteasome. The inhibition of proteasomal activity by the peptidylchloromethanes, Cbz-Leu-Leu-CH2Cl and Cbz-Ala-Ala-Phe-CH2Cl (Cbz, benzyloxycarbonyl), is reversible and predominantly of a competitive type. The enzyme is not activated by any of the compounds that typically stimulate the activities of the eukaryotic proteasome.
