Hodges R S, Zhu B Y, Zhou N E, Mant C T
Department of Biochemistry, University of Alberta, Edmonton, Canada.
J Chromatogr A. 1994 Jul 29;676(1):3-15. doi: 10.1016/0021-9673(94)80452-4.
We have evaluated the potential of reversed-phase liquid chromatography (RPLC) as a probe of hydrophobic interactions involved in protein folding and stability. Our approach was to apply RPLC to a de novo designed model protein system, namely a two-stranded alpha-helical coiled coil. It was shown that the reversed-phase retention behaviour of various synthetic analogues of monomeric alpha-helices and dimeric coiled-coil structures correlated well with their stability in solution, as monitored by circular dichroism during guanidine hydrochloride and temperature denaturation studies. In addition, an explanation is offered as to why amphipathic coiled coils, an important structural motif in many biological systems, are more stable at low pH compared to physiological pH values. The results of this study suggest that not only may RPLC prove to be a useful and rapid complementary technique for understanding protein interactions, but also the de novo designed coiled-coil model described here is an excellent model system for such studies.
我们评估了反相液相色谱（RPLC）作为探究蛋白质折叠和稳定性中疏水相互作用探针的潜力。我们的方法是将RPLC应用于一个从头设计的模型蛋白质系统，即一个双链α-螺旋卷曲螺旋。结果表明，在盐酸胍和温度变性研究过程中，通过圆二色性监测发现，单体α-螺旋和二聚体卷曲螺旋结构的各种合成类似物的反相保留行为与其在溶液中的稳定性密切相关。此外，还解释了为什么两亲性卷曲螺旋作为许多生物系统中的一种重要结构基序，在低pH值下比生理pH值下更稳定。这项研究的结果表明，RPLC不仅可能被证明是一种用于理解蛋白质相互作用的有用且快速的补充技术，而且这里描述的从头设计的卷曲螺旋模型是用于此类研究的优秀模型系统。
