Cooperative folding near the downhill limit determined with amino acid resolution by hydrogen exchange
| Autor: | Tobin R. Sosnick, Wookyung Yu, Karl F. Freed, Isabelle Gagnon, Michael C. Baxa |
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| Rok vydání: | 2016 |
| Předmět: |
Models
Molecular 0301 basic medicine Protein Denaturation Protein Folding 030103 biophysics Magnetic Resonance Spectroscopy Protein Conformation Cooperativity Phi value analysis 03 medical and health sciences Protein structure Computer Simulation Viral Regulatory and Accessory Proteins Denaturation (biochemistry) Folding funnel Amino Acids Multidisciplinary Chemistry Hydrogen Bonding Biological Sciences Contact order Repressor Proteins Kinetics Crystallography 030104 developmental biology Models Chemical Chemical physics Thermodynamics Protein folding Downhill folding Hydrogen |
| Zdroj: | Proceedings of the National Academy of Sciences. 113:4747-4752 |
| ISSN: | 1091-6490 0027-8424 |
| DOI: | 10.1073/pnas.1522500113 |
| Popis: | The relationship between folding cooperativity and downhill, or barrier-free, folding of proteins under highly stabilizing conditions remains an unresolved topic, especially for proteins such as λ-repressor that fold on the microsecond timescale. Under aqueous conditions where downhill folding is most likely to occur, we measure the stability of multiple H bonds, using hydrogen exchange (HX) in a λYA variant that is suggested to be an incipient downhill folder having an extrapolated folding rate constant of 2 × 10(5) s(-1) and a stability of 7.4 kcal·mol(-1) at 298 K. At least one H bond on each of the three largest helices (α1, α3, and α4) breaks during a common unfolding event that reflects global denaturation. The use of HX enables us to both examine folding under highly stabilizing, native-like conditions and probe the pretransition state region for stable species without the need to initiate the folding reaction. The equivalence of the stability determined at zero and high denaturant indicates that any residual denatured state structure minimally affects the stability even under native conditions. Using our ψ analysis method along with mutational ϕ analysis, we find that the three aforementioned helices are all present in the folding transition state. Hence, the free energy surface has a sufficiently high barrier separating the denatured and native states that folding appears cooperative even under extremely stable and fast folding conditions. |
| Databáze: | OpenAIRE |
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