NMR STUDIES OF VACCINIA VIRUS DNA SEQUENCES

痘苗病毒的DNA是一个大的(180kbp)的线性双链 交叉连接的末端，使得在其上形成一个单链圆 变性。终端10 KBP是反向中继器，并且 末端的106个核苷酸形成不完全的碱基配对 发夹；它们参与复制的形成 痘苗病毒的中间体。末端的发夹是非常 富含AT(96%AT)，并包含非对称内部循环和数字 单底凸起。我们对牛痘的结构很感兴趣 含有两个不成对残基(G和C)的病毒DNA片段 在100%富含AT的复式公寓的背景下。我们设计了一个结构 包含这个序列(一个29聚体发夹环DNA)和一个 缺少未配对残基(A)的相应对照构建体 27-聚发夹环DNA)，并获得了一组2D质子核磁共振数据 高分辨率的结构计算。我们已经快完成了 两个分子的质子共振指认，目前我们 对对照的27-聚体序列进行结构计算。 在计算机上开发的交互式计算机图形软件 图形实验室用于精细化的所有阶段：首先， 为了检验实验约束的自我一致性，那么， 监控改进的过程，最后，可视化和 对最终的结构进行分析。

The DNA of vaccinia virus is a large (180 kbp) linear duplex with cross-linked termini, such that a single strand circle forms upon denaturation. The terminal 10 kbp are inverted repeats, and the ultimate 106 nucleotides at each end form incompletely base paired hairpins; they are involved in the formation of replicative intermediates of vaccinia virus. The terminal hairpins are extremely AT-rich (96% AT) and contain an asymmetric internal loop and a number of single base bulges. We are interested in a structure of a vaccinia virus DNA fragment which contains two unpaired residues (G and C) within the context of a 100% AT-rich duplex. We designed a construct containing this sequence (a 29-mer hairpin loop DNA) and a corresponding control construct lacking the unpaired residues (a 27-mer hairpin loop DNA), and acquired sets of 2D proton NMR data for high-resolution structure calculations. We have nearly completed proton resonance assignments for both molecules, and currently we are carrying out structure calculations for the control 27-mer sequence. The interactive computer graphics software developed at the Computer Graphics Laboratory is used at all stages of the refinement: firstly, to examine the self-consistency of the experimental restraints, then, to monitor the course of the refinement, and finally, to visualize and analyze the final structure.