Vu C, Robblee J, Werner K M, Fairman R
Department of Molecular, Cell, and Developmental Biology, Haverford College, 370 Lancaster Ave., Haverford, PA 19041, USA.
Protein Sci. 2001 Mar;10(3):631-7. doi: 10.1110/ps.41101.
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.
了解决定蛋白质稳定性和结构特异性的化学力平衡对于蛋白质设计工作的成功至关重要。具体而言，人们已广泛探索带电荷氨基酸之间的静电相互作用，以了解这种作用力对蛋白质稳定性的贡献。关于静电相互作用作为双链卷曲螺旋中特异性和稳定性决定因素的重要性，已有大量研究，但对于使用此类系统的吸引力大小仍存在重大争议。我们开发了一种四链卷曲螺旋系统，在b和c七肽位置引入了带电荷的残基，以探究电荷相互作用的作用。在此，我们定量测试了不同侧链长度对此类静电相互作用强度的影响。我们合成了在b和c七肽位置均含有天冬氨酸或鸟氨酸的肽，并测试了它们自我缔合以及彼此之间以及与在相同位置含有谷氨酸或赖氨酸的肽进行异源缔合的能力。我们发现，谷氨酸与赖氨酸或鸟氨酸之间的相互作用比涉及天冬氨酸的相应相互作用更有利。在每种情况下，电荷相互作用为卷曲螺旋提供了额外的稳定性，尽管螺旋倾向效应可能在决定这些结构的整体稳定性方面发挥重要作用。
