The effect of poly(ethylene glycol) on the activity, structural conformation and stability of yeast hexokinase

Abstract

In the dilute range (3 mg/L), the effect of poly(ethylene glycol) (PEG) on the enzymatic activity of hexokinase (HK) was investigated by means of spectrophotometry in the absence and presence of PEG400, PEG1500 and PEG4000 in tris(hydroxymethyl)aminomethane–HCl buffer. Comparing the values obtained in pure buffer with those in the polymer solutions, we found that the enzymatic activity increased 6% in the presence of PEG400 and decreased in the range 14–7% in the presence of PEG1500 and PEG4000, respectively. Circular dichroism measurements revealed that in the presence of PEG the α-helix content in HK decreased up to 15% in relation to that measured in pure water. Small-angle X-ray scattering measurements were performed in a higher concentration regime (10 g/L). Data analysis performed from small-angle X-ray scattering curves by means of the intraparticle distance distribution function yielded for HK in water solutions and in PEG400 solutions a radius of gyration, R _g , of (32±1) Å, which is compatible with that obtained from the crystalline dimeric structure of HK. In contrast, in the presence of PEG1500 or PEG4000 large aggregates (without precipitation) were observed. In the presence of glucose and polymer, the aggregation of HK was always favored.

Acknowledgements: The authors thank the LNLS for the use of their facilities and S. Schreier for the use of CD equipment. This work was supported by research grants from FAPESP and CNPq.