Thermodynamic analysis of a designed three-stranded coiled coil
Thermodynamic analysis of a designed three-stranded coiled coil
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DOI：
10.1021/bi961831d
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发表时间：
1996-11-19
期刊：
影响因子：
2.9
通讯作者：
Fairman, R
中科院分区：
文献类型：
--
作者：
Boice, JA;Dieckmann, GR;Fairman, R
螺旋卷曲蛋白质结构基序的研究和成功设计为蛋白质折叠和稳定性的规律提供了更多的洞察力。在这项工作中，我们使用热力学方法来量化控制卷曲线圈的特定齐聚的规则。我们设计了一个高度稳定的三聚体螺旋线圈，通过在七肽重复单元的每个a位放置valine残基，在每个d位放置亮氨酸残基。由沉淀平衡确定该多肽形成一个非常稳定的三聚体，其圆二色光谱的浓度依赖关系遵循协同单体/二聚体/三聚体平衡，二聚体状态为高度不稳定的中间体。通过对数据的分析，确定了该三聚体的三聚化过程的协同性，并提供了该三聚体的稳定自由能为-18.4千卡·摩尔(-1)。通过对在一定浓度下收集的热展开数据的全局分析，测量了热容Delta C-p，在热展开曲线中，Guanidine会诱导冷变性，为Delta C-p提供了一个合理的确定值750卡·度(-1)摩尔(-1)。这相当于每个残基的增量C-p为8.6，称为deg(-1)mol(-1)，与具有明确三级结构的盘绕线圈的预期值很好地对应。
The study and successful design of coiled-coil protein structural motifs have provided much insight into the rules governing protein folding and stability. in this work we use a thermodynamic approach to quantitate the rules that govern the specific oligomerization of coiled coils. We have designed a highly stable trimeric coiled coil by placing valine residues at each a position and leucine residues at each d position of the heptad repeating unit. The peptide forms a very stable trimer as determined by sedimentation equilibrium, and the concentration dependence of its circular dichroism spectrum follows a cooperative monomer/dimer/trimer equilibrium with the dimer state as a highly unstable intermediate, Its guanidinium chloride denaturation curve was collected al several peptide concentrations, and analysis of the data confirms the cooperativity of the trimerization process and provides a free energy of stabilization of -18.4 kcal mol(-1) for the trimer. The heat capacity, Delta C-p, was measured by global analysis of thermal unfolding data collected at a number of guanidinium chloride concentrations, Guanidinium chloride induces cold denaturation in the thermal unfolding curves, providing a reasonably well-determined value for Delta C-p of 750 cal deg(-1) mol(-1). This translates to a Delta C-p of 8.6 cal deg(-1) mol(-1) per residue and corresponds well to that expected of a coiled coil with a well-defined tertiary structure.