Burst-phase expansion of native protein prior to global unfolding in SDS.
Although numerous studies have been directed at understanding early folding events through the characterization of folding intermediates, there are few reports on the very late folding events, i.e. on the events taking place on the native side of the folding barrier and on alternative conformations of the folded state. To shed further light on these issues, we have characterized by protein engineering the structure of an expanded but native-like intermediate that accumulates transiently in the unfolding reaction of the small protein S6 in the presence of SDS. The results show that the SDS micelles attack the native protein in the dead-time of the denaturation experiment, causing an expansion of the hydrophobic core prior to the major unfolding transition. We distinguish two forms of the unfolding intermediate that are correlated with the micellar structure. With spherical micelles, the expansion is seen mainly as a weakening of the interactions which anchor the two alpha-helices to the core of the S6 structure. With cylindrical micelles, prevalent at higher SDS concentrations, the expansion is more global and produces a species which closely resembles the transition-state structure for unfolding in GdmCl. Despite the highly weakened core, the micelle-associated intermediate displays cooperative unfolding, indicating a significant structural plasticity of the species on the native side of the folding barrier in the presence of SDS.
Otzen
Daniel E
DE
Department of Life Sciences, Aalborg University, Sohngaardsholmsvej 49, Aalborg, DK-9000, Sweden.
Oliveberg
Mikael
M
D002942:Circular Dichroism; D003902:Detergents / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D019791:Guanidine / Q000494:pharmacology; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D008823:Micelles; D009154:Mutation / Q000235:genetics; D011489:Protein Denaturation / Q000187:drug effects; D015202:Protein Engineering; D017510:Protein Folding; D017433:Protein Structure, Secondary / Q000187:drug effects; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence; D055672:Static Electricity; D013499:Surface Properties / Q000187:drug effects; D013816:Thermodynamics
D003902:Detergents; D008823:Micelles; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins; D012967:Sodium Dodecyl Sulfate; D019791:Guanidine