PROTEIN STRUCTURE DETERMINATION OF HUMAN Z

RNA编辑酶ADAR 1含有两个同源的左手 Za特异性结合的DNA（Z-DNA）结合结构域Z和Zp 并且对poly（br-dCdG）Z-DNA具有高亲和力（KD = 4 nM）。 获得 对ADARI介导的RNA调节的分子理解 编辑与Z-DNA的识别有关，我们正在确定 Z-DNA结合蛋白结构域Za的高分辨率结构（9.4 kD）。 在500、600和750 MHz下收集光谱。 骨干 从3D 15 N-编辑的光谱获得归属。 TOCSY-HSQC 光谱（35、70和100 ms混合时间）提供了 自旋系统信息，例如几乎所有H，以及许多其他 脂肪族质子 通过3D ′ 5 N-HNHA对H质子进行了归属 江西篇章 通过3D 15顺序分配各个自旋系统 N NOESY-HSQC光谱（70、150和250 ms）。 NOE数据分析 Z有三条P链，称为O链I，II 和三. 连续NOE模式中的两个缺口出现在 脯氨酸残基的位置，其不能被15 N-edited实验 中等NH（i）-NH（i+2）-NOE与 缺乏一个 NH（i）-NH（i+1）-NOE，位于螺旋III和 P-链II表明蛋白质骨架的转向。 分析长 蛋白质骨架中的NOE范围表明， 五个残基。 反平行的P折叠起始于两个残基 螺旋III的C末端，并以C末端终止。 此外，委员会认为， 在P-链I之间检测到NH（i）-NHO）-和H（i-1）-NHO）-NOE。 和Pstrand III，将C-末端反平行P-片层与 P-sheet I.此外， 色氨酸195的芳环质子在P-片层III的质子 亮氨酸176在螺旋III的中心表明， 反平行的P-折叠折叠回到螺旋III上。

The RNA editing enzyme ADAR1 contains two homologous left-handed DNA (Z-DNA) binding domains Z and Zp of which Za binds specifically and with high affinity (KD = 4 nM) to poly(br-dCdG) Z-DNA. To gain a molecular understanding of how the regulation of ADARI-mediated RNA editing relates to the recognition of Z-DNA, we are determining the high resolution structure of the Z-DNA binding protein domain Za (9.4 kD). Spectra were collected at 500, 600 and 750 MHz. Backbone assignments were obtained from 3D 15 N-edited spectra. TOCSY-HSQC spectra (35, 70 and 100 ms mixing times) provided the major part of spin system information, such as nearly all H, and many other aliphatic protons. The H protons were assigned via a 3D '5 N-HNHA spectrum. Individual spin-systems were sequentially assigned by 3D 15 N NOESY-HSQC spectra (70, 150 and 250 ms). Analysis of the NOE data has revealed that Z possesses three P-strands, called 0-strand I, II and III. Two gaps in the sequential NOE pattern arise at the positions of prolines residues, which cannot be detected by 15 N-edited experiments. A medium NH(i)-NH(i+2)-NOE in combination with the lack of a NH(i)-NH(i+l)-NOE at glycine 183 between helix III and P-strand II suggests a turn of the protein backbone. Analysis of long range NOEs in the protein backbone suggests an antiparallel P-sheet of five residues. The antiparallel P-sheet starts two residues C-terminal to helix III and is terminated by the Cterminus. Moreover, a NH(i)-NHO)- and a H,,(i-1)-NHO)-NOE were detected between P-strand I and Pstrand III, connecting the C-terminal antiparallel P-sheet with P-sheet I. Furthermore, well separated long range NOEs between the aromatic ring protons of trytophan 195 in P-sheet III to the protons of leucine 176 in the center of helix III suggests that the antiparallel P-sheet folds back onto helix III.