Synergistic Effect of Charged Amino Acids on the Stability and Fibril Formation of Lysozyme

Osmolytes alter the stability and aggregation of proteins. The effect of mixture of osmolytes is generally not a simple additive of their individual effects. Here, we examine the effect of different mixtures of charged amino acids (AAs) on the stability and fibrillation of lysozyme. Arg, Asp and Glu induce three-state unfolding. The addition of Lys, Asp and Glu increases the thermal stability of lysozyme, whereas Arg does not alter the stability. The mixture of AAs destabilizes the intermediate, but the overall stability of lysozyme is increased. Arg with other AAs in the ratio of 1 : 0.5 and 1 : 1, and the equimolar ratio of Glu+Asp synergistically increases the stability, whereas the other mixtures counteract the stabilizing effects of each other. The apparent time taken for fibrillation is reduced with lower concentrations of Arg (<= 0.75 M) and Glu (0.25 M) and at higher concentrations of Lys (1 M). In the mixed AAs, Arg and Lys with other AAs, and 1 : 1 ratio of Glu+Asp synergistically increases the fibrillation time. These observations suggest that Arg induces more synergistic stabilization and the presence of basic AAs in the mixture increases the fibrillation time compared to acidic AAs which show synergy only at equimolar concentrations. Addition of charged amino acids, Lys, Asp and Glu increases thermal stability of lysozyme. Arg does not alter the stability and induces lag phase during fibrillation. Mixture of amino acids increases overall stability of lysozyme and the mixtures formed with Arg and Lys delayed fibrillation.image