STRUCTURE OF REV1/PCNA COMPLEX IN PEG

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 我们的目标是确定的三维结构的增殖细胞核抗原（PCNA）的DNA聚合酶Rev 1的域的复合物。Rev 1是跨损伤合成DNA聚合酶的一个成员，允许复制越过DNA损伤，使其成为真核生物中DNA损伤诱导突变的关键因素。最近的研究表明Rev 1在DNA损伤处的高持续合成能力使其必须与PCNA相互作用。研究还表明，这种相互作用在DNA损伤反应期间是稳定的。与这些研究一致，我们的体外结合试验，使用量热法和NMR光谱已经证明了PCNA和BRCT域位于N-末端Rev 1之间的紧密相互作用。我们已经获得了适合于结构测定的PCNA/Rev 1-BRCT复合物的晶体。PCNA富集在硒代蛋氨酸中以收集MAD或SAD数据。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Our objective is to determine the three-dimensional structure of the proliferating cell nuclear antigen (PCNA) in complex with a domain from the DNA polymerase Rev1. Rev1, a member of the translesion synthesis DNA polymerases, allows replication past DNA lesions, making it a key factor in DNA damage-induced mutagenesis in eukaryotes. Recent studies have shown that the high processivity of Rev1 at DNA lesions necessitates its interaction with PCNA. It was also shown that this interaction is stabilized during the DNA damage response. In agreement with these studies, our in vitro binding assays using calorimetry and NMR spectroscopy have demonstrated a tight interaction between PCNA and a BRCT domain located at the N-terminus Rev1. We have obtained crystals of the PCNA/Rev1-BRCT complex that are suitable for structure determination. PCNA was enriched in selenomethionine to collect MAD or SAD data.