NMR STRUCTURE OF THE NANOS METAL BINDING DOMAIN

Nanos [nos]是一个后组基因， 果蝇的发育Nanos蛋白发挥其形态发生作用 通过抑制来自母体转录本的蛋白质表达， hunchback [hb]基因两个几乎相同的11个核苷酸重复序列 在hb mRNA的3'非翻译区（3' UTR）中， 纳米效应，这些重复的序列被称为纳米效应。 响应元素（NRE）。nanos C-末端结构域（约120 残基）在双翅目昆虫中高度保守。已经 提出nanos C-末端结构域包含两个CCHC锌结合 识别NRE的基序。我们将描述特异性， 金属与纳米粒子结合的化学计量、亲和性和协同性 C-末端结构域。通过准备各种截短版本的C- 末端结构域，纳米细胞的最小金属和NRE结合序列 将被确定。最小C末端的三级结构 将研究具有最佳金属和RNA结合活性的结构域 使用多维核磁共振（NMR）。基于 纳米C-末端结构域的结构，将假设 通过合作使用体内和体外试验进行突变研究 怀特黑德研究所的鲁思·莱曼教授。这些研究将 为长期目标奠定基础--解决纳米/NRE问题 复杂的结构，以阐明其特定的结构基础， 交互.

Nanos [nos], is a posterior group gene essential for abdominal development in Drosophila. Nanos protein exerts its morphogenic effects by inhibiting protein expression from maternal transcripts of the hunchback [hb] gene. Two nearly identical 11 nucleotide repeat sequences in the 3' untranslated region (3' UTR) of hb mRNA are required for the nanos effect, and these repeated sequences are referred to as nanos response elements (NREs). The nanos C-terminal domain (approximately 120 residues) is highly conserved among Dipteran insects. It has been proposed that the nanos C-terminal domain contains two CCHC zinc binding motifs which recognize NREs. We will characterize the specificity, stoichiometry, affinity and cooperativity of metal binding to the nanos C-terminal domain. By preparing various truncated versions of the C- terminal domain, the minimal metal- and NRE-binding sequence of nanos will be determined. The tertiary structure of the minimal C-terminal domain that has optimal metal and RNA binding activities will be studied using multi-dimensional nuclear magnetic resonance (NMR). Based upon the structure of nanos C-terminal domain, hypotheses will be made for mutational studies using in vivo and in vitro assays by collaboration with Prof. Ruth Lehmann at the Whitehead Institute. These studies will establish the foundation for the long term goal -- solving the nanos/NRE complex structure to elucidate the structural basis for their specific interactions.