Seo Emily S, Sherman John C
Department of Chemistry, 2036 Main Mall, Vancouver, BC, Canada V6T 1Z1.
Biopolymers. 2007;88(5):774-9. doi: 10.1002/bip.20791.
Four-, five-, and six-helix bundle template assembled synthetic proteins (TASPs) have been synthesized using disulfide bonds between cavitand templates and peptides, and characterized in terms of stability and structural specificity. The peptide sequence (CGGGEELLKKLEE LLKKG) used was originally designed for a four-helix bundle. The TASPs were analyzed using CD spectroscopy, chemical denaturation studies, NMR spectroscopy, sedimentation equilibria studies, and hydrophobic dye binding studies to determine the effect of a single peptide sequence when incorporated into bundles with different numbers of helices. If the design was indeed idealized for a four-helix bundle, then the five- and six-helix bundles should be less stable and manifest lower conformational specificity. The TASPs all demonstrated high stability and cooperative unfolding. However, the four-helix bundle was found to be significantly more stable and nativelike compared to the five- and six-helix bundles. This suggests that the peptide sequence is specific to the four-helix bundle, as designed. This result demonstrates the ability to design de novo proteins with specified structure, not just generic stability.
利用穴状配体模板与肽之间的二硫键合成了四螺旋束、五螺旋束和六螺旋束模板组装的合成蛋白（TASPs），并对其稳定性和结构特异性进行了表征。所使用的肽序列（CGGGEELLKKLEE LLKKG）最初是为四螺旋束设计的。通过圆二色光谱（CD光谱）、化学变性研究、核磁共振光谱（NMR光谱）、沉降平衡研究和疏水染料结合研究对TASPs进行分析，以确定当单个肽序列掺入不同螺旋数的束中时的影响。如果该设计确实是针对四螺旋束理想化的，那么五螺旋束和六螺旋束应该稳定性较低且构象特异性较低。所有TASPs均表现出高稳定性和协同解折叠。然而，与五螺旋束和六螺旋束相比，发现四螺旋束的稳定性明显更高且更接近天然状态。这表明肽序列如设计的那样对四螺旋束具有特异性。该结果证明了从头设计具有特定结构而非仅仅是一般稳定性的蛋白质的能力。
