The combined effects of two anti-aggregatory agents, alpha-cyclodextrin and Ca2+, on the refolding process of denatured alpha-amylase.
It is now a well-known fact that alpha-CD (alpha-cyclodextrin) and Ca(2+) are two important folding agents in the re-activation and refolding processes of many denatured enzyme solutions. Bacillus sp. alpha-amylase was extensively denatured in a 6 M guanidinium chloride solution overnight. Under suitable renaturation conditions, 20-30% more activity was recovered in the presence of various concentrations of alpha-CD (0-100 mM). Similarly, 25-30% more activity was recovered under the influence of different concentrations of Ca(2+) (0-100 mM). Regardless of these results, both alpha-CD and Ca(2+) at 100 mM concentrations were separately capable of preventing the extent of aggregate formation by 86 and 77% respectively. These observations clearly indicate that the effects of alpha-CD and/or Ca(2+) on the process of aggregation and refolding (activity recovery) are not synchronized: both agents mostly act as anti-aggregatory agents instead of acting as refolding agents.
Yazdanparast
Razieh
R
Institute of Biochemistry and Biophysics, P.O. Box 13145-1384, University of Tehran, Tehran, Iran. yazdan@ibb.ut.ac.ir
Khodarahmi
Reza
R
D002118:Calcium / Q000494:pharmacology*; D002122:Calcium Chloride / Q000737:chemistry; D002413:Cations, Divalent / Q000494:pharmacology; D019791:Guanidine / Q000737:chemistry; D011489:Protein Denaturation; D017510:Protein Folding; D000516:alpha-Amylases / Q000737:chemistry* / Q000378:metabolism; D047391:alpha-Cyclodextrins / Q000494:pharmacology*
D002413:Cations, Divalent; D047391:alpha-Cyclodextrins; D000516:alpha-Amylases; D019791:Guanidine; D002122:Calcium Chloride; D002118:Calcium; C032613:alpha-cyclodextrin