Fig. 5.

(A) The cp6-^SFGFP-5/6-ssrA protein (10 μM; NCBI accession code JF951870) was cleaved with thrombin and incubated with ClpXP (1 μM ClpX₆; 2 μM ClpP₁₄), 4 mM ATP, and an ATP-regeneration system. ClpXP extraction of the terminal β strand resulted in time-dependent loss of 467-nm fluorescence and 400-nm fluorescence (data not shown). The initial rate of this reaction (0.2 min⁻¹ enz⁻¹) was slow, but within error of the rate of ClpXP degradation of uncleaved cp6-^SFGFP-5/6-ssrA (data not shown). (B) Absorbance spectra of thrombin-cleaved cp6-^SFGFP-5/6-ssrA (closed triangles), the cp6-^SFGFP-5 protein after ClpXP strand extraction and purification by S200 gel filtration (open triangles), and cp6-^SFGFP-ssrA denatured by incubation at pH 2 (squares).