Figure 8.

Dilution experiment in the presence of an α-casein trap to determine whether ClpAP can degrade substrates in a single cycle of substrate binding and product release. Substrate–ClpA–ClpP complexes were formed by incubating 0.8 pmol each of ClpA and ClpP with 0.9 pmol of [³H]RepA or [³H]α-casein in 5 μl of buffer A containing 1 mM ATP[γ-S] for 10 min at 23°C. Complexes were diluted to 1 ml with buffer A containing 4 μM α-casein, 0.005% Triton X-100, and 1 mM ATP, and incubated an additional 10 min at 23°C. Reaction products were precipitated with 20% TCA, and radioactivity in the acid-soluble fractions was measured. Control experiments are as shown.