Ubiquitin folds through a highly polarized transition state.
The small alpha/beta protein ubiquitin has been used as a model system for experimental and computational studies on protein folding for many years. Here, we present a comprehensive phi-value analysis and characterize the structure and energetics of the transition state ensemble (TSE). Twenty-seven non-disruptive mutations are made throughout the structure and a range of phi-values from zero to one are observed. The values cluster such that medium and high values and found only in the N-terminal region of the protein, whilst the C-terminal region has consistently low phi-values. In the TSE, the main alpha-helix appears to be fully formed (two phi-values which specifically probe helical structure are one) and the helix is stabilized by packing against the first beta-turn, which is partially structured. In striking comparison, the phi-values in the C-terminal region are all very low, suggesting that this region of the protein is largely unstructured in the TSE. Data are consistent with a nucleation-condensation mechanism in which there is a highly polarized folding nucleus comprising the first beta-hairpin and the alpha-helix. Data presented from the protein engineering study and phi-value analysis are compared with results from other experimental studies and also computational studies.
Went
Heather M
HM
Chemistry Department, Centre for Protein Engineering, Lensfield Road, Cambridge CB2 1EW, UK.
Jackson
Sophie E
SE
D007700:Kinetics; D008958:Models, Molecular; D016296:Mutagenesis; D017354:Point Mutation; D015202:Protein Engineering; D017510:Protein Folding; D017433:Protein Structure, Secondary; D013816:Thermodynamics; D025801:Ubiquitin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D025801:Ubiquitin