Open AccessRefolding and Reactivation of Liver Alcohol Dehydrogenase after Dissociation and Denaturation in 6M Guanidine Hydrochloride
Author(s) -
GERSCHITZ Johan,
RUDOLPH Rainer,
JAENICKE Rainer
Publication year - 1978
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1978.tb12411.x
Subject(s) - guanidine , chemistry , kinetics , dissociation (chemistry) , alcohol dehydrogenase , nad+ kinase , hydrochloride , denaturation (fissile materials) , cofactor , enzyme , bivalent (engine) , biophysics , chromatography , biochemistry , stereochemistry , organic chemistry , nuclear chemistry , biology , physics , quantum mechanics , metal
Horse‐liver alcohol dehydrogenase has been dissociated and denatured by 6 M gaunidinium hydrochloride. Removal of the denaturat under optimum conditions of the solvent leads to partial reactivation. The concentrations of the enzyme, as well as the coenzyme (NAD + and Zn 2+ , affect the reactivation significantly, since high concentrations promote the formation of inactive aggregation products. Analyzing the kinetics of reactivation and reassociation, conditions far from equilibrium of dissociation‐association provide maximum yieldddd (∼ 70%). The sigmoidal kineic traces suggest a superposition of first‐order transconformation and‐second‐order association reactions; the latter are corrobosrated by the concentration dependence of thereactivation reaction. The coenzyme, NAD + , had no the kinetics of reactivation. Addition of Zn 2+ leads to a significant decrease of the rate and yield of ractivation. The process of renaturation, as reflected by the regain of navive fluorescence show complex kinetics: rapid relaxations are followed by slower first‐order and secon‐order processes which parallel reactivtion.