Increased thermal stability of proteins in the presence of amino acids

Abstract

This study is a systematic attempt to understand the roles of osmolytes in protecting proteins against denaturing stress. Thermal denaturation of cytochrome c has been studied in the presence of various concentrations of all L-amino acids that are more hydrophobic than glycine and have a solubility of 0.1 M or higher in water at 25 degrees C. The basic observations are as follows. (1) Arginine and histidine destabilize the native protein; both Tm (the midpoint of thermal transition) and delta GDH2O (25 degrees C) (the Gibbs energy of stabilization) decrease with increasing amino acid concentration. (2) Isoleucine, leucine and phenylalanine have no effect on Tm and deltaGDH2O (25 degrees C). (3) Valine and less hydrophobic amino acids stabilize the protein in terms of Tm but deltaGDH2O (25 degrees C) is unchanged. This observation was confirmed by the study of isothermal denaturation of cytochrome c by guanidinium chloride which suggested that delta GDH2O is independent of osmolyte concentration, but Cm (the midpoint of transition) is increased in their presence. (4) In the case of stabilizers, change in Tm/mol of amino acid decreases with increasing hydrophobicity of these osmolytes.

Effect of Arginine on Glycation and Stability of Recombinant Human Interferon-Gamma

“…Arginine is a widely used additive in biotechnology and pharmaceutical industry for suppressing aggregation, assisting in refolding of denatured proteins, and enhancing protein solubility (2,20,21,22). Although many hypotheses have been proposed to explain the protective effect of arginine against protein aggregation (1,3,12), the exact mechanism of this phenomenon is still obscure.…”

Control of Globular Protein Thermal Stability in Aqueous Formulations by the Positively Charged Amino Acid Excipients

“…2 Interestingly, usually featured in lists of commonly used excipients are the three positively charged amino acids: arginine, lysine and histidine. 1,3 Many studies have shown lysine to stabilise proteins in solution [4][5][6][7] despite the fact that it has been demonstrated to perturb enzyme function. Quite often 35 lysine is shown to stabilise proteins to a much lesser extent than other amino acids and small molecules.…”

“…The data for the effects of histidine on protein stability are much more scarce and much less 40 uniform, with certain studies showing a destabilising effect on proteins 5 and others showing histidine as a protein stabiliser. 7,9 No clear mechanism has yet been suggested for the effect of histidine on protein stability.…”

Promotion and Suppression of Thermal Aggregation of β-Lactoglobulin by Arginine: A Concentration Dependent Mechanism

“…The stability of the native state of dissolved proteins is generally not affected by the presence of L-arginine. Earlier it was reported that L-arginine has been found to destabilize RNaseA [13] and Cytochrome C [14] slightly. On the other hand, Reddy et al (2005) hardly found any destabilizing effect of L-arginine.HCl up to concentrations level of 1.0 M in the case of hen egg white Lysozyme [15].…”