ИСТИНА

In vivo proteins exist in molecular crowding conditions, i.e. when free volume is significantly limited. In vitro such conditions are simulated by concentrated solutions of “inert” polymers (crowders or crowding agents). It was shown that such environment can affect proteins folding, structural dynamics and functional activity. Here, the effect of one of crowders, polyethyleneglycol (PEG) on the folding/unfolding of D-glucose/D-galactose-binding protein (GGBP) was studied. It was shown that PEG 12 and 4 kDa at high concentrations promote the increase of GGBP secondary structure content and shift of GGBP denaturation curves to higher concentrations of chemical denaturant guanidine hydrochloride (GdnHCl). The dependences of GGBP molar ellipticity at 222 nm on GdnHCl concentrations have local minimum near 2 M GdnHCl in these PEG solutions. It may indicate the existence of intermediate state of GGBP with higher content of secondary structure in comparison with the unfolded protein in these GdnHCl concentrations. It was established that denaturation and renaturation curves of GGBP in 12 and 4 kDa PEG solutions with concentrations 300 and 200 mg/ml do not coincide. It was shown that in solutions with low GdnHCl concentrations CD spectra of GGBP after renaturation in the presence of PEG 12 and 4 kDa at high concentrations significantly differ from CD spectra obtained after denaturation of protein in same conditions. All obtained data suggest that PEG of high molecular mass (12 and 4 kDa) at high concentrations (300 and 200 mg/ml) promote the change of protein free energy landscape and folding/unfolding pathway of GGBP. This work was supported by a grant from Russian Science Foundation RSCF 14-24-00131