Sticky-end assembly of a designed peptide fiber provides insight into protein fibrillogenesis
Sticky-end assembly of a designed peptide fiber provides insight into protein fibrillogenesis
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DOI：
10.1021/bi000246g
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发表时间：
2000-08-01
期刊：
影响因子：
2.9
通讯作者：
Woolfson, DN
中科院分区：
文献类型：
--
作者：
Pandya, MJ;Spooner, GM;Woolfson, DN
卷曲螺旋基序为研究分子自组装提供了简单的系统。我们设计了两个28个残基的肽组装成一个扩展的卷曲螺旋纤维。核心和侧翼离子对中的互补相互作用用于指导交错的异二聚体。这些具有“粘性末端”以促进长纤维的形成。为了比较，我们还合成了一种肽的排列形式，以与另一种肽结合，并形成具有不能纵向结合的“平端”的典型异二聚体。使用圆二色性光谱在溶液中监测这两对的组装。在每种情况下，混合肽导致在222 MI处增加的浓度依赖性圆二色性信号，与所需的α-螺旋结构一致。对于所设计的产生纤维的肽混合物，我们还观察到在流动取向过程中的线性二色性效应，表明存在长纤维结构。部分对准样品的X射线纤维衍射保存指示卷曲螺旋结构的图案。此外，我们使用电子显微镜直接观察纤维形成。有趣的是，观察到的纤维至少几百微米长，比预期的二聚体卷曲螺旋设计厚20倍。这一额外的厚度意味着设计结构的横向关联。我们提出，互补功能存在于重复结构的类型，我们描述促进横向组装，并在某些自然系统中，一个类似的机制可能是fibrillogenesis。
Coiled-coil motifs provide simple systems for studying molecular self-assembly. We designed two 28-residue peptides to assemble into an extended coiled-coil fiber. Complementary interactions in the core and flanking ion-pairs were used to direct staggered heterodimers. These had "sticky-ends" to promote the formation of long fibers. For comparison, we also synthesized a permuted version of one peptide to associate with the other peptide and form canonical heterodimers with "blunt-ends" that could not associate longitudinally. The assembly of both pairs was monitored in solution using circular dichroism spectroscopy. In each case, mixing the peptides led to increased and concentration-dependent circular dichroism signals at 222 MI, consistent with the desired alpha-helical structures. For the designed fiber-producing peptide mixture, we also observed a linear dichroism effect during flow orientation, indicative of the presence of long fibrous structures. X-ray fiber diffraction of partially aligned samples Save patterns indicative of coiled-coil structure. Furthermore, we used electron microscopy to visualize fiber formation directly. Interestingly, the fibers observed were at least several hundred micrometers long and 20 times thicker than expected for the dimeric coiled-coil design. This additional thickness implied lateral association of the designed structures. We propose that complementary features present in repeating structures of the type we describe promote lateral assembly, and that a similar mechanism may underlie fibrillogenesis in certain natural systems.