BPTI folding pathways
BPTI has three disulfides (30-51, 5-55 and 14-38). Reduction of
the DS's thus gives 6 CysSH. Reoxidation is performed with a
mixture of oxidized and reduced glutathione, GSSG and GSH,
under alkaline conditions. Creighton used rapid quenching with
acid or excess iodoacetate to trap the remaining free SH groups,
and separate the resulting labeled protein by chromatographic
methods. The major findings were that there was one predominant
one-disulfide intermediate, out of the 15 possible ones, which
contained the native 30-51 DS. All subsequent intermediates
contained this DS. What was especially striking about Creighton's
results was that the subsequent two-disulfide species favored two
non-native second disulfide bonds. This led to idea of trapped
misfolded states. However, this view has been challenged in BPTI