MOLECULAR STUDIES OF PROTEIN-DNA INTERACTIONS

我们试图在分子水平上了解生物体在适应遗传多样性需要的同时保持其基因组完整性的各种方式。 我们的研究工作目前集中在两个相关的过程，同源重组和DNA错配修复。错配修复以E.大肠杆菌甲基指导的错配修复途径，目标是通过DNA复制错误，同源重组和自发DNA损伤引起的碱基对错配。 大肠杆菌mutS和mutL基因编码的细菌错配修复系统的组成部分。大肠杆菌中，在原核生物和真核生物中高度保守。 编码错配修复酶的人类基因缺陷与散发性和遗传性癌症有关。我们有兴趣了解MutS蛋白错配识别的分子机制。 我们已经确定了MutS的二聚化结构域映射到一个高度保守的螺旋-U-转角-螺旋区的羧基末端的蛋白质。我们的研究表明，MutS的二聚化是必不可少的错配修复在体内和体外的DNA结合和ATP水解。 Taq MutS和一个MutS错配复合物的晶体结构已被解决。 这些结构揭示了一种诱导的错配识别机制，涉及一个复合错配结合位点，存在一个以前不受重视的复合ATP酶活性位点和一个连接这两个功能域的递质区域。

We seek to understand at a molecular level the various ways by which an organism maintains the integrity of its genome while accommodating the need for genetic diversity. Our research efforts currently focus on two related processes, homologous recombination and DNA mismatch repair. Mismatch repair, exemplified by the E. coli methyl-directed mismatch repair pathway, targets base pair mismatches that arise through DNA replication errors, homologous recombination and spontaneous DNA damage. Components of the bacterial mismatch repair system encoded by the mutS and mutL genes in E. coli, are highly conserved in both prokayotes and eukaryotes. Defects in human genes encoding mismatch repair enzymes have been implicated in sporadic and hereditary cancers. We are interested in understanding the molecular mechanisms involved in mismatch recognition by the MutS protein. We have identified a dimerization domain of MutS that maps to a highly conserved helix-U-turn-helix region at the carboxy terminus of the protein. Our studies indicate that dimerization of MutS is essential for mismatch repair in vivo and for DNA binding and ATP hydrolysis in vitro. The crystal structures of Taq MutS and a MutS-mismatch complex have been solved. The structures reveal an induced-fit mechanism of mismatch recognition involving a composite mismatch binding site, the existence of a previously unappreciated composite ATPase active site and a transmitter region connecting these two functional domains.