Figure 3.

GroEL residues that can interact with rhodanese in the cage. (A) Solvent-accessible surface of the GroES-associated GroEL ring in the GroEL–GroES–(ADP)₇ complex (PDB code 1AON) viewed from outside, inside, close-up to the GroEL/GroES-binding site from inside and above. GroEL and GroES subunits are coloured in green and orange, respectively. The residues mutated to cysteine are labelled and coloured (hydrophilic, cyan; acidic, red; basic, blue; hydrophobic, yellow). The putative polypeptide-binding site is depicted by a red circle. (B) Cross-linking between rhodanese_His6 (Rho) and cysteine-introduced SR398 mutants. Diamide was added to form disulphide bonds before (ATP−) or at 0.1 min after the addition of GroES and ATP (ATP+). Non-reducing SDS–PAGE gels were stained with CBB. (C, D) Time course of the cross-linking between rhodanese_His6 and SR398(R231C) (C) or SR398(V264C) (D) during the assisted folding. After the folding reaction was started, aliquots were oxidized by diamide at indicated times. Time courses of cross-linked products (x-link, closed squares) and free rhodanese_His6 (free, open squares), and rhodanese activity (closed circles) are plotted.