STUDIES ON THE ACTIVATION BY ATP OF THE 26-S PROTEASOME COMPLEX FROM RAT SKELETAL-MUSCLE

The 26 S proteasome complex is thought to catalyse the breakdown of ub iquitinated proteins within eukaryotic cells. In addition it has been found that the complex also degrades short-lived proteins such as orni thine decarboxylase in a ubiquitin-independent manner. Both proteolyti c processes are paralleled by the hydrolysis of ATP. Here we show that ATP also affects the hydrolytic activity towards fluorigenic peptide substrates by the 26 S proteasome complex from rat skeletal muscle tis sue. Low concentrations of ATP (about 25 mu M) optimally activate the so-called chymotryptic and tryptic activity by increasing the rate of peptide hydrolysis but not peptidylglutamylpeptide hydrolysis. Activat ion of the enzyme by ATP is transient but this effect can be enhanced and prolonged by including in the assay an ATP-regenerating system, in dicating that ATP is hydrolysed by the 26 S proteasome complex. Althou gh ATP cannot be substituted for by adenosine 5'-[beta,gamma-methylene ]triphosphate or AMP, hydrolysis of the phosphoanhydride band of ATP s eems not to be necessary for the activation process of the proteasome complex, a conclusion drawn from the findings that ATP analogues such as adenosine 5'-[beta,gamma-imido]triphosphate, adenosine 5'-O-[gamma- thio]triphosphate, adenosine 5'-O-[beta-thio]diphosphate and adenosine 5'-[alpha,beta-methylene]triphosphate give the same effect as ATP, an d vanadate does not prevent ATP activation. These effects are independ ent of the presence of Mg2+. Thus, ATP and other nucleotides may act a s allosteric activators of peptide-hydrolysing activities of the 26 S proteasome complex as has also been found with the ion protease from E scherichia coli.