DESIGNED DNA CRYSTALS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 我们的研究计划旨在利用基于分支DNA的新型高分子构建块，作为专门设计的晶体排列3D结构基元的基础。最终目标是提供能够结合、定向和并列各种分子的大分子支架，从生物大分子到有机导体和光存储元件。我们提出并成功地确定了这样一个设计的三维系统的结构-张拉整体三角形。这种结构是一个稳健的基序，具有三重旋转主干对称性，由三个向线性独立方向的螺旋组成，即它们的螺旋轴并不都共享同一平面。螺旋通过三个像Holliday一样的交叉点成对连接，从而产生一个交替的上下主题。最近，我们报道了[郑等人]。自然461，74-77(2009)]X射线晶体结构为一个张拉整体三角形，包含单一分子物种，由三个螺旋结构域组成，每个螺旋结构域包含两个双螺旋转角。每个三角形都以菱面体的一个顶点为中心，形成一个大的空腔。我们最近的努力指向了几个不同的方向，所有这些都是基于已公布的两转三角形的结构。(A)我们已成功地将三角形的螺旋转数从2个增加到3个和4个，并确定了它们的晶体结构。虽然精细结构都具有从设计中预测的结构参数，但所获得的晶体的分辨率随着螺旋转数的增加而降低。我们正在探索改变连接三角形的粘性末端长度的影响，以及使用天然DNA而不是合成DNA对晶体分辨率的影响。(B)我们正在尝试将客体分子结合到晶体结构的内腔中；三种不同长度边(2、3和4圈)的结构的空腔分别为~100 Nm~3、~375 Nm~3和~1000 Nm~3。客体种类从蛋白质、多肽和多肽仿制物到染料、金属纳米颗粒和DNA片段。通过共结晶法和浸渍法制得了这类配合物的类晶。一份描述DNA晶体晶体结构的报告已经准备好提交，该报告包含两个不同的三角形，根据设计进行结晶。此外，我们还证明了晶体的颜色可以通过染料分子与不同分子的共价结合来控制。通常，我们的晶体是弱衍射的，表现出高度的马赛克，并经常出现在空间群P1中。因此，这种晶体需要较长的曝光时间、较小的摆动角以及360度以上的最佳数据收集。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Our research program is designed to utilize new types of macromolecular building blocks based on branched DNA, as the basis of specifically designed crystalline arrangements 3D structural motifs. The ultimate goals are to provide macromolecular scaffoldings, capable of binding, orienting and juxtaposing a variety of molecules, from biological macromolecules to organic conductors and optical memory components. We proposed and succeeded in determining structures of such a designed 3D system, the tensegrity triangle. This structure is a robust motif with three-fold rotational backbone symmetry, consisting of three helices that are directed in linearly independent directions, i.e., their helix axes do not all share the same plane. The helices are connected pair-wise by three Holliday-like crossover points, so as to produce an alternating over-and-under motif. Recently, we reported [Zheng et al. Nature 461, 74-77 (2009)] the X-ray crystal structure to 4 ¿ of a tensegrity triangle containing a single molecular species, comprised of three helical domains, each containing two double helical turns. Each triangle is centered on a vertex of a rhombohedron, creating a large cavity. Our more recent efforts have been directed in several different directions, all based on the structure of the published two-turn triangle. (A) We have been successful in increasing the numbers of helical turns in the triangle from 2 to 3 and 4 and in determining their crystal structures. While the refined structures all have the structural parameters predicted from their designs, the resolutions of the crystals obtained decrease with the increase in the number of helical turns. We are exploring the effect of changing the length of sticky ends that link the triangles, and also the impact of using natural DNA, rather than synthetic DNA, on the resolution of the crystals. (B) We are attempting to incorporate guest molecules into the internal cavities of the crystal structures; the cavities of the three structures with different length edges (2, 3 and 4 turns) are ~100 nm3, ~375 nm3 and ~1000 nm3. The guest species range from proteins, peptides, and peptidomimetics to dyes, metallic nanoparticles and segments of DNA. Isomorphous crystals have been obtained of such complexes both by co-crystallization and soaking. A report describing the crystal structure of a DNA crystal containing two distinct triangles programmed to crystallize according to the design is ready for submission. In addition, we have also demonstrated that the colors of the crystals can be controlled by the covalent attachment of dye molecules to the different molecules. As a rule our crystals are weakly diffracting, display a high degree of mosaicity and frequently appear in space group P1. Consequently the crystals require long exposure times, small oscillation angles and optimally data collection over 360 degrees.