THE PROTEASOME ACTIVATOR 11-S-REGULATOR OR PA28 - CONTRIBUTION BY BOTH ALPHA-SUBUNIT AND BETA-SUBUNIT TO PROTEASOME ACTIVATION

The proteasome 11 S regulator (REG) consists of two homologous subunit s, REG alpha and REG beta. Each subunit is capable of activating the p roteasome, and when combined, they form superactive REG alpha/REG beta complexes. We have previously shown that a highly conserved loop in t he REG alpha crystal structure is critical for proteasome activation. We now show that hetero-oligomers formed from REG alpha activation loo p mutants and wild-type REG beta or vice versa are partially active. B y contrast, heterooligomers bearing mutations in the activation loops of REG alpha and REG beta subunits are inactive, demonstrating that bo th alpha and beta subunits contribute to proteasome activation. We hav e also characterized REG proteins with mutations near or at their C te rmini. Partially active REG alpha(Y249C) and REG alpha(M247V) and an i nactive REG alpha subunit bearing five additional C-terminal amino aci ds formed active hetero-oligomers with REG beta. REG beta subunits lac king 1, 2, or 9 C-terminal amino acids did not bind or activate the pr oteasome, but each of these mutants formed partially active hetero-oli gomers with the monomer REG alpha(N50Y). However, hetero-oligomers for med from REG subunits lacking the last 14 amino acids were unable to b ind the proteasome. Thus, C-terminal regions of both alpha and beta su bunits are required for hetero-oligomers to bind the proteasome.