Designing heterodimeric two-stranded α-helical coiled-coils -: Effects of hydrophobicity and α-helical propensity on protein folding, stability, and specificity
Designing heterodimeric two-stranded α-helical coiled-coils -: Effects of hydrophobicity and α-helical propensity on protein folding, stability, and specificity
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DOI：
10.1074/jbc.m204257200
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发表时间：
2002-10-04
影响因子：
4.8
通讯作者：
Hodges, RS
中科院分区：
文献类型：
--
作者：
Litowski, JR;Hodges, RS
E/K大肠杆菌是一种异二聚体大肠杆菌，已被设计为一种通用的肽捕获和传递系统，用于生物传感器和表达和亲和纯化标签等应用。在这个设计中，异二聚体的形成是通过在七重体的e和g位置放置带电残基来指定的，这样大肠杆菌在这些位置上包含所有的谷氨酸残基，而大肠杆菌在这些位置上包含所有的赖氨酸残基。E/K大肠杆菌的亲和力和稳定性已被修改，以允许更大范围的关联和解离条件。用异亮氨酸代替缬氨酸增加大肠杆菌核心的疏水性，稳定性分别提高了2.81和3.73 kcal/mol （0.47 kcal/mol / replacement）。以丙氨酸取代丝氨酸，增加核心外残基的α -螺旋倾向，稳定性分别提高2.68和3.28 kcal/mol（0.41和0.45 kcal/mol/取代）。这些序列的改变产生了一系列异二聚体大肠杆菌，其稳定性在6.8 ~ 11.2 kcal/mol之间，极大地扩展了它们在蛋白质工程和生物医学领域的应用范围。
The E/K coli, a heterodimeric colied-coli, has been designed as a universal peptide capture and delivery system for use in applications such as biosensors and as an expression and affinity purification tag. In this design, heterodimer formation is specified through the placement of charged residues at the e and g positions of the heptad repeat such that the E coli contains all glutamic acid residues at these positions, and the K coli contains all lysine residues at these positions. The affinity and stability of the E/K coli have been modified to allow a greater range of conditions for association and dissociation. Increasing the hydrophobicity of the colied-coli core, by substituting isoleucine for valine, gave increases in stability of 2.81 and 3.73 kcal/mol (0.47 kcal/ mol/substitution). Increasing the alpha-helical propensity of residues outside the core, by substituting alanine for serine, yielded increases in stability of 2.68 and 3.28 kcal/mol (0.41 and 0.45 kcal/mol/substitution). These sequence changes yielded a series of heterodimeric colied-colis whose stabilities varied from 6.8 to 11.2 kcal/mol, greatly expanding their scope for use in protein engineering and biomedical applications.