Arginine hydrochloride enhances the dynamics of subunit assembly and the chaperone-like activity of α-crystallin

Abstract

Purpose: α-Crystallin, a major eye lens protein, bears homology with small heat shock proteins (sHsps) and exhibits molecular chaperone-like activity. Structural perturbation by temperature or low concentrations of denaturants leads to enhancement of its chaperone-like activity. We have earlier demonstrated similar enhancement of chaperone-like activity using biologically compatible solutes such as arginine hydrochloride and aminoguanidine. The purpose of the present study is to get an insight into the mechanism of the arginine induced enhancement of chaperone-like activity of α-crystallin. Methods: The effect of arginine hydrochloride on the chaperone-like activity of α-crystallin at 25 °C was studied using DTT induced aggregation of insulin as a model system. Changes in the accessibility of the thiol group near the end of the α-crystallin domain in the absence and the presence of arginine hydrochloride were studied using dithiobisnitrobenzoic acid. Fluorescence resonance energy transfer studies were performed to investigate changes in the dynamics of the subunit assembly. Urea induced denaturation studies of α-crystallin were carried out to investigate structural destabilization of α-crystallin, if any, in the presence of arginine hydrochloride. Results: Arginine hydrochloride increases the chaperone-like activity of α-crystallin several fold towards DTT induced aggregation of insulin at room temperature. Our study shows that both the extent and the rate of accessibility of the thiol group are increased in the presence of arginine. Fluorescence resonance energy transfer experiments show that arginine hydrochloride significantly increases the subunit exchange between the oligomers of α -crystallin. Arginine induced structural perturbation and loosening of subunit assembly of α-crystallin leads to overall destabilization of the protein as reflected by the urea denaturation study. Conclusions: Arginine perturbs the tertiary and quaternary structure of α-crystallin and enhances the dynamics of the subunit assembly leading to enhanced chaperone-like activity. Thus, in addition to size, surface hydrophobicity, and charge distribution, the dynamics of the subunit assembly appears to be one of the critical factors that can modulate the chaperone activity.