Effects of charged amino acids at b and c heptad positions on specificity and stability of four‐chain coiled coils

Abstract

An understanding of the balance of chemical forces responsible for protein stability and specificity of structure is essential for the success of efforts in protein design. Specifically, electrostatic interactions between charged amino acids have been explored extensively to understand the contribution of this force to protein stability. Much research on the importance of electrostatic interactions as specificity and stability determinants in two‐stranded coiled coils has been done, but there remains significant controversy about the magnitude of the attractive forces using such systems. We have developed a four‐stranded coiled‐coil system with charged residues incorporated at b and c heptad positions to explore the role of charge interactions. Here, we test quantitatively the effects of varying sidechain length on the magnitude of such electrostatic interactions. We synthesized peptides containing either aspartate or ornithine at both b and c heptad positions and tested their ability to self‐associate and to hetero‐associate with one another and with peptides containing glutamate or lysine at the same positions. We find that interactions between glutamate and either lysine or ornithine are more favorable than the corresponding interactions involving aspartate. In each case, charged interactions provide additional stability to coiled coils, although helix propensity effects may play a significant role in determining the overall stability of these structures.