Folding kinetics of the all-beta-sheet protein human basic fibroblast growth factor, a structural homolog of interleukin-1beta.
The refolding and unfolding kinetics of the all-beta-sheet protein human basic fibroblast growth factor (hFGF-2) were studied by fluorescence spectroscopy. The kinetics of the unfolding transition are monophasic. The refolding reaction at high and low guanidinium chloride (GdmCl) concentrations is best described by mono- and biphasic folding, respectively. Refolding and unfolding of hFGF-2 (155 amino acids) is very slow compared with other non-disulfide-bonded monomeric proteins of similar size. For example, the rate constant for unfolding at 4.5 mol.liter(-1) GdmCl is 0.006 s(-1), and the refolding rate constants at 0.4 mol.liter(-1) GdmCl are 0.01 s(-1) and 0.0009 s(-1) (15 degrees C, pH 7.0). A characterization of the thermodynamic nature of the folding process using transition state theory revealed that the slow refolding is almost exclusively controlled by entropic factors, namely the strong loss of conformational freedom during refolding. The rate of the slow unfolding kinetics is mainly (and at low denaturant concentrations exclusively) controlled by the large positive change in enthalpy. hFGF-2 shows similar slow folding kinetics to that of its structural homolog interleukin-1beta. Since both proteins show very little sequence identity, it is suggested that their slow folding kinetics are determined by the complex beta-sheet arrangement of the native molecules.
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GBF National Research Center for Biotechnology, Biochemical Engineering Division, Mascheroder Weg 1, 38124 Braunschweig, Germany.
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D016222:Fibroblast Growth Factor 2 / Q000737:chemistry* / Q000187:drug effects; D019791:Guanidine / Q000494:pharmacology; D006801:Humans; D007375:Interleukin-1 / Q000737:chemistry*; D007700:Kinetics; D017510:Protein Folding; D017433:Protein Structure, Secondary; D013696:Temperature; D013816:Thermodynamics
D007375:Interleukin-1; D016222:Fibroblast Growth Factor 2; D019791:Guanidine