Figure 3.

Isolation of RepGFP-ClpA complexes by gel filtration. [³H]RepGFP (160 pmol) was incubated with 200 pmol of ClpA for 15 min at 24°C in 100-μl reaction mixtures containing buffer B and 1 mM ATP[γS]. The mixtures were then applied to a 0.7 × 15 cm column of Sephacryl S200 HR at 24°C equilibrated with 20 mM Tris⋅HCl, pH 7.5/10% (vol/vol) glycerol/100 mM KCl/5 mM DTT/0.1 mM EDTA/5 mM MgCl₂/0.5 mM ATP[γS]. Fractions (150 μl) were collected into microtiter plates. Relative fluorescence (blue circles) and radioactivity (open black squares) were measured in aliquots of the fractions.