Rapid formation of the native 14-38 disulfide bond in the early stages of BPTI folding.
Using recombinant variants of BPTI, we have determined the rate constants corresponding to formation of each of the fifteen possible disulfide bonds in BPTI, starting from the reduced, unfolded protein. The 14-38 disulfide forms faster than any of the other 14 possible disulfides. This faster rate results from significantly higher intrinsic chemical reactivities of Cys-14 and Cys-38, in addition to local structure in the reduced protein that facilitates formation of the 14-38 disulfide bond. This disulfide bond is found in native BPTI. Our results suggest that a significant flux of folding BPTI molecules proceed through the one-disulfide intermediate with the 14-38 disulfide bond, denoted [14-38], that has recently been detected on the BPTI folding pathway. In addition to providing a detailed picture of the early events in the folding of BPTI, our results address quantitatively the effect of local structure in the unfolded state on folding kinetics.
Dadlez
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Howard Hughes Medical Institute, Whitehead Institute for Biomedical Research, Department of Biology, Massachusetts Institute of Technology, Cambridge 02142, USA.
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D000595:Amino Acid Sequence; D000818:Animals; D007611:Aprotinin / Q000096:biosynthesis / Q000737:chemistry*; D002417:Cattle; D003001:Cloning, Molecular; D003545:Cysteine; D004220:Disulfides; D004926:Escherichia coli; D005978:Glutathione / Q000031:analogs & derivatives; D019803:Glutathione Disulfide; D007202:Indicators and Reagents; D007461:Iodoacetates; D019807:Iodoacetic Acid; D007700:Kinetics; D008956:Models, Chemical; D008961:Models, Structural; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D014508:Urea
D004220:Disulfides; D007202:Indicators and Reagents; D007461:Iodoacetates; D010446:Peptide Fragments; D011994:Recombinant Proteins; D014508:Urea; D007611:Aprotinin; D005978:Glutathione; D003545:Cysteine; D019803:Glutathione Disulfide; D019807:Iodoacetic Acid