Mo X, Bailin T, Sadofsky M J
Institute of Molecular Medicine and Genetics, Medical College of Georgia, Augusta, Georgia 30912-2650, USA.
J Biol Chem. 1999 Mar 12;274(11):7025-31. doi: 10.1074/jbc.274.11.7025.
An essential step in the development of the vertebrate immune system is the DNA level rearrangement of the antigen receptor genes. This process, termed "V(D)J recombination," begins with DNA cleavage at the appropriate sites mediated by the two proteins RAG1 and RAG2. We report here that the two proteins cooperate to bind DNA with significantly higher specificity than either protein alone. Gel purification of the triple complex is performed in the absence of any cross-linking agents. Both proteins remain present in the complex, and UV cross-linking using iodouridine-containing probes shows that RAG1 makes close contacts in both the heptamer and nonamer motifs. The two proteins are also shown to associate with each other in the absence of any DNA. These findings refine our understanding of the protein-DNA interactions that accompany cleavage at the recombination signals.
脊椎动物免疫系统发育过程中的一个关键步骤是抗原受体基因的DNA水平重排。这一过程被称为“V(D)J重组”，始于由RAG1和RAG2这两种蛋白质介导的在适当位点的DNA切割。我们在此报告，这两种蛋白质协同作用结合DNA，其特异性显著高于单独的任何一种蛋白质。在没有任何交联剂的情况下对三元复合物进行凝胶纯化。两种蛋白质都保留在复合物中，使用含碘尿苷的探针进行紫外线交联表明，RAG1在七聚体和九聚体基序中都有紧密接触。还表明这两种蛋白质在没有任何DNA的情况下也会相互结合。这些发现完善了我们对重组信号处切割所伴随的蛋白质-DNA相互作用的理解。
