Department of Chemistry and Biochemistry, University of Arizona, Tucson, AZ 85721-0088, USA.
Protein Eng Des Sel. 2011 Sep;24(9):765-71. doi: 10.1093/protein/gzr027. Epub 2011 Jun 14.
Regions of amino-acid sequence that are compatible with multiple folds may facilitate evolutionary transitions in protein structure. In a previous study, we described a heuristically designed chameleon sequence (SASF1, structurally ambivalent sequence fragment 1) that could adopt either of two naturally occurring conformations (α-helical or β-sheet) when incorporated as part of the C-terminal dimerization subdomain of two structurally divergent transcription factors, P22 Cro and λ Cro. Here we describe longer chameleon designs (SASF2 and SASF3) that in the case of SASF3 correspond to the full C-terminal half of the ordered region of a P22 Cro/λ Cro sequence alignment (residues 34-57). P22-SASF2 and λ(WDD)-SASF2 show moderate thermal stability in denaturation curves monitored by circular dichroism (T(m) values of 46 and 55°C, respectively), while P22-SASF3 and λ(WDD)-SASF3 have somewhat reduced stability (T(m) values of 33 and 49°C, respectively). (13)C and (1)H NMR secondary chemical shift analysis confirms two C-terminal α-helices for P22-SASF2 (residues 36-45 and 54-57) and two C-terminal β-strands for λ(WDD)-SASF2 (residues 40-45 and 50-52), corresponding to secondary structure locations in the two parent sequences. Backbone relaxation data show that both chameleon sequences have a relatively well-ordered structure. Comparisons of (15)N-(1)H correlation spectra for SASF2 and SASF3-containing proteins strongly suggest that SASF3 retains the chameleonism of SASF2. Both Cro C-terminal conformations can be encoded in a single sequence, showing the plausibility of linking different Cro folds by smooth evolutionary transitions. The N-terminal subdomain, though largely conserved in structure, also exerts an important contextual influence on the structure of the C-terminal region.
氨基酸序列中与多种折叠相容的区域可能促进蛋白质结构的进化转变。在之前的研究中，我们描述了一个启发式设计的变色龙序列（SASF1，结构不稳定序列片段 1），当作为两个结构上不同的转录因子 P22 Cro 和 λ Cro 的 C 末端二聚化亚结构域的一部分被掺入时，它可以采用两种天然构象（α-螺旋或β-折叠）。在这里，我们描述了更长的变色龙设计（SASF2 和 SASF3），在 SASF3 的情况下，它对应于 P22 Cro/λ Cro 序列比对中有序区域的完整 C 末端半区（残基 34-57）。P22-SASF2 和 λ（WDD）-SASF2 在圆二色性（CD）监测的变性曲线中表现出中等的热稳定性（T m 值分别为 46 和 55°C），而 P22-SASF3 和 λ（WDD）-SASF3 的稳定性略有降低（T m 值分别为 33 和 49°C）。（13）C 和（1）H NMR 二级化学位移分析证实 P22-SASF2（残基 36-45 和 54-57）具有两个 C 末端α-螺旋，λ（WDD）-SASF2（残基 40-45 和 50-52）具有两个 C 末端β-折叠，这与两个亲本序列中的二级结构位置相对应。骨架弛豫数据表明，两个变色龙序列都具有相对有序的结构。对含有 SASF2 和 SASF3 的蛋白质的（15）N-（1）H 相关光谱的比较强烈表明，SASF3 保留了 SASF2 的变色龙性质。两种 Cro C 末端构象都可以在单个序列中编码，这表明通过平滑的进化转变将不同的 Cro 折叠连接起来是合理的。尽管结构上基本保守，N 末端亚结构域也对 C 末端区域的结构施加了重要的上下文影响。
