Disulfide bonds BPTI refolding

Disulfide Bonds - Bonds between cysteine residues in a protein help to stabilize it once it has folded. Bovine pancreatic trypsin inhibitor (BPTI), which has 3 disulfide bonds in its 58 amino acid sequence, is one of the stablest proteins known. When the bonds are in place, it can be denatured at lOO C only in very acid solutions. Removing one disulfide bond reduces its resistance to thermal denaturation considerably. Removing all of the disulfide bonds causes it to unfold at room temperature. If time is allowed for refolding and the disulfide bonds are reformed, virtually 100% of the original activity of BPTI can be recovered, indicating that the sequence of amino acids contains enough information to properly reestablish the folding of the polypeptide. 

Bovine pancreatic trypsin inhibitor (BPTI), a small protein with 56 amino acid residues (Fig. 7), is the first one for which a detailed map of the refolding pathways was deciphered. The native state of BPTI contains three disulfide (S-S) bonds formed between six Cys residues. Native state is specified by [30-51 5-55 14—38] bonds. This notation indicates that Cys forms an S-S bond with Cys, and so on. Reduction of the S-S bonds unfolds BPTI. By using S-S bond formation as a progress variable, Creighton [79-83] devised ingenious methods to trap the disulfide-bonded intermediates along the folding pathway. 

Fig. 9.14. Competition between I]-BPTI and reduced intermediates and refolded forms of BPTI for anti-N-antibodies (A) (0)i native ( ), refolded (A), reduced-carboxymethylated BPTI (B) (A) single-disulfide bond intermediate (5-30) and (A)> (30-51) (C) (O) two-disulfide bond intermediate (30-51,14-38) and (A), the mixture of (30-51,5-14) and (30-51,5-38) and (D) (A) nativelike two-disulfide bond species (30-51, 5-55) and (A) N cm and (O) N gJS (from Creighton et al., 1978, courtesy of Creighton).

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