ENZYMATIC AND STRUCTURAL SIMILARITIES BETWEEN THE ESCHERICHIA-COLI ATP-DEPENDENT PROTEASES, CLPXP AND CLPAP

Escherichia coli ClpX, a member of the Clp family of ATPases, has ATP- dependent chaperone activity and is required for specific ATP-dependen t proteolytic activities expressed by ClpP, Gel filtration and electro n microscopy showed that ClpX subunits (M-r 46,000) associate to form a six-membered ring (M-r similar to 280,000) that is stabilized by bin ding of ATP or nonhydrolyzable analogs of ATP, ClpP, which is composed of two seven-membered rings stacked face-to-face, interacts with the nucleotide-stabilized hexamer of ClpX to form a complex that could be isolated by gel filtration. Electron micrographs of negatively stained ClpXP preparations showed side views of 1:1 and 2:1 ClpXP complexes i n which ClpP was flanked on either one or both sides by a ring of ClpX , Thus, as was seen for ClpAP, a symmetry mismatch exists in the bondi ng interactions between the seven-membered rings of ClpP and the six-m embered rings of ClpX, Competition studies showed that ClpA may have a slightly higher affinity (similar to 2-fold) for binding to ClpP, Mix ed complexes of ClpA, ClpX, and ClpP with the two ATPases bound simult aneously to opposite faces of a single ClpP molecule were seen by elec tron microscopy, In the presence of ATP or nonhydrolyzable analogs of ATP, ClpXP had nearly the same activity as ClpAP against oligopeptide substrates (>10,000 min(-1)/tetradecamer of ClpP), Thus, ClpX and ClpA interactions with ClpP result in structurally analogous complexes and induce similar conformational changes that affect the accessibility a nd the catalytic efficiency of ClpP active sites.