Evolutionary conservation and variation of protein folding pathways. Two protease inhibitor homologues from black mamba venom.
The pathways of unfolding and refolding of three homologous proteins are shown to be closely related. This implies that folding pathways, as well as the final folded conformation, have been largely conserved during the presumed evolutionary divergence of these proteins from a common ancestor. The pathways of the homologous proteins I and K from black mamba venom were determined here, using the disulphide interaction between their six cysteine residues to trap and identify the intermediate states, and are compared with those determined previously in the same way for the homologous bovine pancreatic trypsin inhibitor. The major one- and two-disulphide intermediates are the same with all three proteins; their kinetic roles are similar, although there are differences in the rates at which they are interconverted and in the minor intermediates that accumulate. As a consequence, different pathways may predominate with another homologous protein, even though the various most favourable pathways are the same. The energetics of the folding transitions and the stabilities of the folded states differ substantially for the three proteins. The differences in stabilities of the fully folded states are primarily reflected kinetically in the rate-determining rearrangements of the native-like conformation; the rates and equilibria of the other steps are not affected markedly. With the less stable proteins, the direct folding pathway of sequential formation of the three correct disulphide bonds becomes significant and is the most facile when considered on a solely intramolecular basis.
D000596:Amino Acids / Q000032:analysis; D007611:Aprotinin; D005075:Biological Evolution; D002852:Chromatography, Ion Exchange; D004220:Disulfides; D004546:Elapid Venoms; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D011480:Protease Inhibitors; D011487:Protein Conformation
D000596:Amino Acids; D004220:Disulfides; D004546:Elapid Venoms; D011480:Protease Inhibitors; C034915:toxin I, black mamba venom; C038787:toxin K, black mamba venom; D007611:Aprotinin