Pii: S1357-2725(01)00003-6
نویسندگان
چکیده
The course of refolding and reactivation of urea-denatured creatine kinase (ATP; creatine N-phosphotransferase, EC 2.7.3.2) has been studied in the absence and presence of molecular chaperonin GroEL. The enzyme was denatured in Tris–HCl buffer containing 6 M urea for 1 h. In the refolding studies, the denatured enzyme was diluted 60-fold into the same buffer containing GroEL or not for activity, turbidity, fluorescence measurements and polyacrylamide gel electrophoresis. The results show that the reactivation process is dependent of creatine kinase concentration in the concentration range 2.5–4 mM. The levels of activity recovery decrease with increasing enzyme concentration because of the formation of wrong aggregates. The molecular chaperonin GroEL can bind the refolding intermediate of creatine kinase and thus prevent the formation of wrong aggregates. This intermediate is an inactive dimeric form that is in a conformation resembling the ‘molten globule’ state. © 2001 Elsevier Science Ltd. All rights reserved.
منابع مشابه
Pii: S1357-2725(01)00048-6
Aggregation of 3 M guanidine hydrochloride denatured creatine kinase (ATP: creatine N-phosphotransferase, EC 2.7.3.2) occurs after dilution into the refolding solution. Proline, glycerol and heparin sodium act as folding aids which can effectively inhibit aggregation of creatine kinase during refolding. Proline at 1 M concentration, glycerol at 10% concentration and heparin at 25 mg/ml not only...
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