Kinetic roles and conformational properties of the non-native two-disulphide intermediates in the refolding of bovine pancreatic trypsin inhibitor.
The most productive folding pathway of reduced bovine pancreatic trypsin inhibitor (BPTI) proceeds through the disulphide intermediates (30-51), (30-51, 5-14), and (30-51, 5-38); these are important kinetic intermediates in folding, even though the latter pair contain non-native disulphide bonds. Analogues of these intermediates have been prepared by protein engineering methods and their conformational properties examined by circular dichroism and 1H-nuclear magnetic resonance. The (30-51), (30-51, 5-14) and (30-51, 5-38) analogues exhibit comparable degrees of stable structure, which cannot include those portions of the polypeptide chain involving Cys5, Cys14 and Cys38. These properties are consistent with the roles of (30-51, 5-14) and (30-51, 5-38) in the folding pathway of BPTI, which demand that they exhibit a considerable degree of conformational flexibility in part of the molecule.
Darby
N J
NJ
MRC Laboratory of Molecular Biology, Cambridge, England.
van Mierlo
C P
CP
Scott
G H
GH
Neuhaus
D
D
Creighton
T E
TE
D000818:Animals; D007611:Aprotinin / Q000737:chemistry / Q000648:ultrastructure*; D002417:Cattle; D002942:Circular Dichroism; D004220:Disulfides; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D011487:Protein Conformation; D011489:Protein Denaturation; D013056:Spectrophotometry, Ultraviolet
D004220:Disulfides; D007611:Aprotinin