NMR STUDY OF DNA/RNA WITH INVERTED POLARITY/ANOMERIC CT

癌基因c-erbB-2的高表达与 几种肿瘤的恶性细胞转化和预后不良 例如乳腺癌和卵巢癌以及腺癌和 可能在星形细胞脑瘤中。这个项目的总体目标是 是为了了解它的结构和热力学参数 新型反义c_erbB-2反义寡核苷酸 反常中心和极性反转。我们计划开展一项 体系结构和热力学性质的系统研究 下面描述的系统为了能够预测和 优化β异常DNA延伸的位置和长度， α异构体中的修饰碱基和糖残基 用于最佳稳定性、核酸酶抗性和核糖核酸酶的成分 H活动，这将导致设计出强大的第二代 反义剂。使用的主要工具是核磁共振波谱， 由UV和CD光谱和核酸酶敏感性支持 学习。 1)α-异构体ODN/RNA双链的研究与设计 激活核糖核酸酶H.长度和要求的测定 贝塔异常延伸，两侧是阿尔法异常组件与 核糖核酸酶H活性及DNA外切酶和内切酶抗性。 2)DNA/DNA解聚体溶液结构的测定 含有α-异构体核苷酸和间隔区的双链 3‘-3’磷酸二酯键。阿尔法无规碱基是 以平行的方式嵌入整个反平行螺旋中。 每个底座的两侧都有一个不同寻常的3英尺3英寸和5英尺5英寸的连杆。这个 每一种双链的稳定性将通过紫外线来确定 融化研究。 3)DNA/RNA双链溶液结构的核磁共振分析 DNA链是否含有α-异常核苷酸和 位于3‘-3’磷酸二酯连接处的间隔基。每条DNA链都会 含有单一的阿尔法异构体碱基，以便清楚地区分 在单个碱基替换中。每一个的稳定性 这些双链将使用UV熔融轮廓来确定。 4)提高了ODN/RNA的热力学稳定性 混血儿。将探索几种化学修饰，以进一步 改善ODN与RNA的结合。 5)第三代ODN的设计、性能和结构。 利用从目标1、3和4获得的结果，更有效 具有优越热力学和核糖核酸酶H的抗c-erbB-2寡核苷酸 激活属性。核磁共振将被用来验证我们的设计 策略。

Increased expression of the oncogene c-erbB-2 is associated with malignant cell transformation and poor prognosis in several tumors such as breast and ovarian cancer and in adenocarcinoma and possibly in astrocytic brain tumors. The overall goal of this project is to understand the structural and thermodynamic parameters of novel c_erbB-2 antisense oligonucleotides that contain inverted anomeric centers and polarity reversals. We plan to carry out a systematic study of the structural and thermodynamic properties of the systems described below in order to be able to predict and optimize, position and length of beta anomeric DNA stretches, modified bases and sugar residues in the alpha anomeric components, for optimal stability, nuclease resistance, and Rnase H activity, that will result in the design of potent second generation antisense agents. The main tool used is NMR spectroscopy, supported by UV and CD spectroscopy and nuclease sensitivity studies. 1) Study and design of alpha anomeric ODN/RNA duplexes which activate Rnase H. Determination of the length and requirement of a beta anomeric stretch flanked by alpha anomeric components vs. Rnase H activity nd DNA exo- and endonuclease resistance. 2) Determination of the solution structure of DNA/DNA decamer duplexes that contain alpha anomeric nucleotides and a spacer at the 3'-3' phosphodiester linkage. Alpha anomeric bases are embedded in a parallel manner in an overall antiparallel helix. Each base is flanked by an unusual 3'3' and 5'5' linkage. The stability of each of these duplexes will be determined by UV melting studies. 3) NMR analysis of the solution structure of DNA/RNA duplexes were the DNA strand contains alpha anomeric nucleotides and spacers at the 3'-3' phosphodiester linkage. Each DNA strand will contain a single alpha anomeric base in order to clearly distinguish among the individual base substitutions. The stability of each of these duplexes will be determined using UV melting profiles. 4) Improvement of the thermodynamic stability of the ODN/RNA hybrids. Several chemical modifications will be explored to further imrpove the ODN-RNA binding. 5) Design, properties and structure of third generation ODNs. Utilizing the results obtained from Aims 1,3 and 4, more potent anti-c-erbB-2 ODNs with superior thermodynamic and Rnase H activation properties. NMR will be used to verify our design strategy.