We describe the guanidinium hydrochloride induced folding kinetics of the four-helix-bundle protein Rop wild-type (wt) under equilibrium conditions at three temperatures. The choice of appropriate denaturant conditions inside the transition range permitted, in combination with equilibrium transition curves, the determination of both unfolding and refolding rate constants. The ratio of the rate constants at zero denaturant concentration provided equilibrium constants and standard free energy changes that are in good agreement with values obtained in previous differential scanning calorimetry studies. The DeltaG0D values for 19, 25 and 40 degrees C calculated from the present kinetic studies are, respectively, 66.8, 70.8 and 57.2 kJ.mol-1. The unfolding reactions are extremely slow under these conditions. Equilibrium was reached only after 18, 12 and 6 days at 19, 25 and 40 degrees C. These results demonstrate that for Rop wt high stability correlates with slow folding kinetics. Study holds ProTherm entries: 15091, 15092, 15093 Extra Details: Additive:EDTA 1mM Rop protein; folding kinetics; slow equilibrium folding; stability
Submitter: Connie Wang
Submission Date: April 24, 2018, 8:45 p.m.
|Number of data points||9|
|Proteins||Regulatory protein rop ; Regulatory protein rop|
|Assays/Quantities/Protocols||Experimental Assay: Cm temp:40.0 C ; Experimental Assay: m temp:40.0 C ; Experimental Assay: dG_H2O temp:40.0 C ; Experimental Assay: Cm temp:25.0 C ; Experimental Assay: m temp:25.0 C ; Experimental Assay: dG_H2O temp:25.0 C ; Experimental Assay: Cm temp:19.0 C ; Experimental Assay: m temp:19.0 C ; Experimental Assay: dG_H2O temp:19.0 C|
|Libraries||Mutations for sequence MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHADELYRSCLARFGDDGENL|