The enzyme rhodanese can be reactivated after denaturation in guanidinium chloride.

摘要: For the first time, enzyme rhodanese has been refolded after denaturation in guanidinium chloride (GdmHCl). Renaturation was by either (a) direct dilution into assay, (b) intermediate buffer, or (c) dialysis followed concentration and centrifugation. Method preferentially retained active whose specific activity 1140 IU/mg, which fell to 898 IU/mg 6 days. The of native is 710 IU/mg. Progress curves were linear for dialyzed enzyme, kinetic analysis showed it had same Km thiosulfate as but apparently displayed a higher turnover number. denatured directly diluted assay mix many three phases: lag during no product formed; order reactivation; an steady state. An induction period determined extrapolating steady-state line time axis. percent reactivation 7% (t1/2 = 10 min) increased between GdmHCl start independent presence thiosulfate. period, decreased zero incubation increased, There observable differences renatured protein electrophoresis fluorescence spectroscopy. Previous reports some refolding urea-denatured (Stellwagen, E. (1979) J. Mol. Biol. 135, 217-229) confirmed, extended, compared with results using GdmHCl. A working hypothesis that involves intermediates partition inactive products. These may result from nucleation two domains, exposes hydrophobic surfaces become interdomain interface correctly folded protein.