ATP BINDING, BUT NOT ITS HYDROLYSIS, IS REQUIRED FOR ASSEMBLY AND PROTEOLYTIC ACTIVITY OF THE HSLVU PROTEASE IN ESCHERICHIA-COLI

HslVU is an ATP-dependent protease consisting of two multimeric compon ents: the HslU ATPase and the HslV peptidase. To gain an insight into the role of ATP hydrolysis in protein breakdown, we determined the ins ulin B-chain-degrading activity and assembly of HslVU in the presence of ATP and its nonhydrolyzable analogs. While beta,gamma-methylene-ATP could not support the proteolytic activity, beta,gamma-imido-ATP supp orted it to an extent less than 10% of that seen with ATP. Surprisingl y, however, HslVU degraded insulin B-chain even more rapidly in the pr esence of ATP gamma S than with ATP. Furthermore, the ability of ATP a nd its analogs in supporting the proteolytic activity was closely corr elated with their ability in supporting the oligomerization of HslU an d the formation of the HslVU complex. However, ADP, which is capable o f supporting the HslU oligomerization, could not support the HslVU com plex formation or the proteolytic activity, suggesting that the confor mation of the ADP-bound HslU oligomer is different from that of ATP-bo und form. Thus, it appears that ATP-binding, but not its hydrolysis, i s essential for assembly and proteolytic activity of HslVU. (C) 1997 A cademic Press.