The underlying mechanism for the diversity of disulfide folding pathways.
The disulfide folding pathway of bovine pancreatic trypsin inhibitor (BPTI) is characterized by the predominance of folding intermediates with native-like structures. Our laboratory has recently analyzed the folding pathway(s) of four 3-disulfide-containing proteins, including hirudin, potato carboxypeptidase inhibitor, epidermal growth factor, and tick anticoagulant peptide. Their folding mechanism(s) differ from that of BPTI by 1) a higher degree of heterogeneity of 1- and 2-disulfide intermediates and 2) the presence of 3-disulfide scrambled isomers as folding intermediates. To search for the underlying causes of these diversities, we conducted kinetic analyses of the reductive unfolding of these five proteins. The experiment of reductive unfolding was designed to evaluate the relative stability and interdependence of disulfide bonds in the native protein. It is demonstrated here that among these five proteins, there exists a striking correlation between the mechanism(s) of reductive unfolding and that of oxidative folding. Those proteins with their native disulfide bonds reduced in a collective and simultaneous manner exhibit both a high degree of heterogeneity of folding intermediates and the accumulation of scrambled isomers along the folding pathway. A sequential reduction of the native disulfide bonds is associated with the presence of predominant intermediates with native- like structures.
Chang
J Y
JY
Research Center for Protein Chemistry, Institute of Molecular Medicine, University of Texas, Houston, Texas 77030, USA. rchang@imm2.imm.uth.tmc.edu
Li
L
L
Bulychev
A
A
D007611:Aprotinin / Q000378:metabolism; D060829:Arthropod Proteins; D004220:Disulfides / Q000378:metabolism*; D004229:Dithiothreitol / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000378:metabolism; D006629:Hirudins / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins; D007700:Kinetics; D008956:Models, Chemical; D008958:Models, Molecular; D010084:Oxidation-Reduction; D010455:Peptides / Q000378:metabolism; D010940:Plant Proteins / Q000378:metabolism; D011480:Protease Inhibitors; D011489:Protein Denaturation; D017510:Protein Folding; D017434:Protein Structure, Tertiary
D060829:Arthropod Proteins; D004220:Disulfides; D006629:Hirudins; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides; D010940:Plant Proteins; D011480:Protease Inhibitors; C063967:tick anticoagulant peptide; D004815:Epidermal Growth Factor; D007611:Aprotinin; D004229:Dithiothreitol