Effects of guanidine hydrochloride on the conformation and enzyme activity of streptomycin adenylyltransferase monitored by circular dichroism and fluorescence spectroscopy.

摘要: Equilibrium denaturation of streptomycin adenylyltransferase (SMATase) has been studied by CD spectroscopy, fluorescence emission and binding the hydrophobic dye 1-anilino-8-naphthalene sulfonic acid (ANS). Far-UV spectra show retention 90% native-like secondary structure at 0.5 M guanidine hydrochloride (GdnHCl). The mean residue ellipticities 222 nm enzyme activity plotted against GdnHCl concentration showed loss about 50 75% 35 60% 0.75 1.5 GdnHCl, respectively. At 6 there was leading to formation GdnHCl-induced unfolded state as evidenced spectroscopy well measuring enzymatic activity. denaturant-mediated decrease in intensity 5 red shift λmax point gradual unfolding SMATase when is added up from a maximum M. Decreasing ANS (∼5 nm) were observed this compared native folded state, indicating partial destruction surface patches protein molecule on denaturation. Disruption disulfide bonds resulted sharp hydrophobicity protein, that are held even denatured state. Acrylamide potassium iodide quenching intrinsic tryptophan conformation with majority residues exposed solvent, 20% them negatively charged environment.