Effects of ammonium sulfate on the unfolding and refolding of the variable and constant fragments of an immunoglobulin light chain.
The equilibria and kinetics of unfolding and refolding by guanidine hydrochloride of the VL and CL fragments of a type kappa immunoglobulin light chain were studied in the presence of ammonium sulfate using circular dichroism and tryptophyl fluorescence at pH 7.5 and 25 degrees C. The unfolding equilibria of the VL and CL fragments were described in terms of the two-state transition. The midpoints of unfolding in the absence of ammonium sulfate were at 0.9 and 1.2 M guanidine hydrochloride for the CL and VL fragments respectively. The transition curves were shifted to higher concentrations of guanidine hydrochloride by 1.4 and 1.6 M for the CL and VL fragments, respectively, per mole of ammonium sulfate. Unfolding reactions of the VL and CL fragments in 3 M guanidine hydrochloride followed first-order kinetics, and the rate constants for the two proteins were both greatly decreased by the presence of ammonium sulfate. The refolding reaction of the CL fragment in 0.3 M guanidine hydrochloride consisted of two phases, and the rate constants were increased a little by the presence of ammonium sulfate. The refolding reaction of the VL fragment in 0.3 M guanidine hydrochloride followed first-order kinetics, and the rate was not affected by the presence of ammonium sulfate. These results showed that ammonium sulfate stabilizes the CL and VL fragments mainly by decreasing the unfolding rate.
D000645:Ammonium Sulfate / Q000494:pharmacology*; D002942:Circular Dichroism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007127:Immunoglobulin Constant Regions; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D007700:Kinetics; D011487:Protein Conformation / Q000187:drug effects; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics
D007127:Immunoglobulin Constant Regions; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D000645:Ammonium Sulfate