Paradoxical acceleration of dithiothreitol-induced aggregation of insulin in the presence of a chaperone.

Natalya Artemova,Bella Gurvits,Boris Kurganov,Zoya Bumagina,Konstantin Muranov

doi:10.3390/ijms11114556

Abstract

The kinetics of dithiothreitol (DTT)-induced aggregation of human recombinant insulin and the effect of α-crystallin, a representative of the family of small heat shock proteins, on the aggregation process have been studied using dynamic light scattering technique. Analysis of the distribution of the particles by size measured in the course of aggregation showed that the initial stage of the aggregation process was the stage of formation of the start aggregates with a hydrodynamic radius (Rh) of about 90 nm. When studying the effect of α-crystallin on the rate of DTT-induced aggregation of insulin, it was demonstrated that low concentrations of α-crystallin dramatically accelerated the aggregation process, whereas high concentrations of α-crystallin suppressed insulin aggregation. In the present study, at the molar stoichiometric ratio (insulin:α-crystallin) less than 1:0.5, a pronounced accelerating effect of α-crystallin was observed; whereas a ratio exceeding the value of 1:0.6 caused suppression of insulin aggregation. The mechanisms underlying the dual effect of α-crystallin have been proposed. It is assumed that heterogeneous nucleation occurring on the surface of the α-crystallin particle plays the key role in the paradoxical acceleration of insulin aggregation by α-crystallin that may provide an alternative biologically significant pathway of the aggregation process.

Highlights

The kinetics of dithiothreitol (DTT)-induced aggregation of human recombinant insulin and the effect of -crystallin, a representative of the family of small heat shock proteins, on the aggregation process have been studied using dynamic light scattering technique
Studies of the protective role of chaperone-like proteins that prevent insulin aggregation may be important for neuroendocrinology, as many pathogenetic problems of a neurodegenerative nature and several other diseases are associated with dysfunctions of insulin [5]
DTT-induced aggregation of insulin was studied by dynamic light scattering (DLS) at 25 C in 50 mM sodium phosphate buffer, pH 7.0, containing 0.15 M NaCl at various concentrations of the protein

Summary

Introduction

The kinetics of dithiothreitol (DTT)-induced aggregation of human recombinant insulin and the effect of -crystallin, a representative of the family of small heat shock proteins, on the aggregation process have been studied using dynamic light scattering technique. In the present work we investigated the effect of -crystallin on the kinetics of DTT-induced aggregation of human recombinant insulin in a wide range of the -crystallin concentrations. This choice was determined by the following considerations: On one hand, the development of modern concepts of protein folding and aggregation has resulted in the discovery of numerous nonpathogenic proteins and peptides, having the ability to form amyloid-like structures, which can induce conformational diseases [4]. Studies of the protective role of chaperone-like proteins that prevent insulin aggregation may be important for neuroendocrinology, as many pathogenetic problems of a neurodegenerative nature and several other diseases are associated with dysfunctions of insulin [5]

Results

Discussion

Conclusion