Abstract The normal CH 2 CH 2 SCH 3 side-chain of Met52 of bovine pancreatic trypsin inhibitor was converted to CH 2 CH 2 S + (CH 3 ) 2 with methyl iodide. After unfolding and breakage of the three disulphide bonds, the reduced protein refolded some three to six times more slowly than unmodified bovine pancreatic trypsin inhibitor. This was shown to be due to the decreased occurrence of the normal one- and two-disulphide initial intermediates. Modifications of the Met52 side-chain suggest that it normally has an important conformational role in the initial stages of folding, participating in extensive hydrophobic interactions with other parts of the protein. This role is different from that in the final folded state, where modification produced no detectable change in stability.