Characterization of SBP1 Binding Determinant

初步证据表明果蝇的结构 结合蛋白1（sbp 1）基因是一个很好的候选基因， 参与DNA重组、复制和/或修复。 的 spb 1基因是一个单拷贝基因， 在组织和生命阶段中富集的转录物 快速细胞分裂。 SPB 1：β-半乳糖苷酶融合蛋白 优先结合序列中的三链DNA结构 独立的方式，并似乎拥有链置换 活动从核酸序列预测的蛋白质， sbp 1 cDNA序列与原核DNA序列有一定的相似性 连接酶。 该提案的主要目的是确定，在单一的- 核苷酸分辨率，特异性结合决定簇存在于 优先识别的三链DNA结构， 由SBP 1绑定。 这将首先通过化学方法实现 探测孤立的结构， DNA的结构。 探针可触及的区域为 预测为单链或扭曲的双链 DNA中的区域。 随后，测定了 优选结合底物的修饰形式的SBP 1将 进行分析。 导致减少或取消的修改 结合的重要性将表明SPB 1的决定因素 约束力 %%% 交换，或交叉，是一个事件， 基因在减数分裂期间在染色体之间交换。 的 这一过程涉及两个DNA螺旋的断裂， 交易所 人们普遍认为，不寻常的DNA结构是 在重组过程中形成。 该提案旨在 了解这些不寻常的DNA结构是如何被识别的， 解决了

Preliminary evidence indicates that the Drosophila Structure Binding Protein 1 (sbp1) gene is an excellent candidate for involvement in DNA recombination, replication, and/or repair. The spb1 gene is a single copy gene producing three different transcripts that are enriched in tissues and life stages involved in rapid cell division. The SPB1:beta-galatosidase fusion protein preferentially binds three-stranded DNA structures in a sequence independent manner, and appears to possess a strand displacement activity. The protein predicted from the nucleic acid sequence of sbp1 cDNAs shares some sequences similarity with prokaryotic DNA ligases. The primary objective of this proposal is to define, at single- nucleotide resolution, the specific binding determinant present in three-stranded DNA structures that is preferentially recognized and bound by SBP1. This will be accomplished by first chemically probing isolated structures to model the three dimensional structure of the DNA. Regions that are accessible to the probe are predicted to be either single stranded or distorted double stranded regions in DNA. Subsequently, the relative binding affinity of SBP1 for modified versions of the preferred binding substrate will be assayed. Modifications resulting in a reduction or elimination of binding will indicate the importance of a determinant for SPB1 binding. %%% Recombination, or crossing over, is the event in which blocks of genes are exchanged between chromosomes during meiosis. The process involves breakage of two DNA helices and the subsequent exchange. It is generally accepted that unusual DNA structures are formed during recombination. This proposal is directed toward understanding how these unusual DNA structures are recognized and resolved.