Genetic Recombination by E. coli's RecBCD Pathway

同源染色体通过“广义基因重组”相互交换片段。这一过程在细菌病毒lambda中尤其容易实现，因为它的物理简单性和我们对该病毒的深入了解。我们的工作已经证实，在lambda中影响重组的四条已识别的酶途径中的每一条都受到DNA双链断裂的刺激，并提出了所有自发的普遍重组都依赖于这种断裂的观点。在所有的途径中，这些因素所起的作用并不相同。在其中三条途径(“类红色途径”)中，Break直接参与重组事件，而在另一条途径(RecBCD)中，Break充当酶的进入位点，该酶在离Break任意距离处被特定核苷酸序列(CHI)激活后，沿着DNA移动，实现重组。对于这两种类型的路径，都提供了高度定义的模型。这笔拨款旨在测试这些型号和它们的替代版本。一个统一的模型假设，在两种类型的途径中，在距离双链断裂任意距离的双链之间建立了同源配对。在类似红色的途径中，这些配对区域致力于通过将切割分子的一条链从切割区域降解到配对区域而进行重组。在RecBCD途径中，在切入点进入的RecBCD酶在不重组的情况下分解这些区域，除非酶由于传递CHI或类似CHI的序列而丢失了D亚基。

Homologous chromosomes exchange segments with each other via "generalized genetic recombination." This process is especially accessible in the bacterial virus lambda because of its physical simplicity and our intimate knowledge of that virus. Our work has established that each of the four identified enzymatic pathways that effect recombination in lambda is stimulated by DNA double-chain breaks and suggests the view that all spontaneous generalized recombination is dependent upon such breaks. The role played by these is not the same in all the pathways. In three of the pathways ("Red-like pathways") the breaks participate directly in the recombination event while in the other pathway (RecBCD) the break acts as an entry site for an enzyme that moves along the DNA effecting recombination after it is activated by a specific nucleotide sequence (Chi) at an arbitrary distance from the break. For both types of pathway highly defined models have been offered. This grant seeks to test those models and alternate versions of them. A unifying model supposes that in both types of pathway homologous pairing is established between duplexes at arbitrary distance from the double chain-break. In Red-like pathways, these regions of pairing are committed to recombination by degradation of one chain of the cut molecule from the cut to the region of pairing. In the RecBCD pathway, the RecBCD enzyme, which enters at the cut, resolves these regions without recombination, unless the enzyme has lost the D subunit as a result of passing Chi or a Chi-like sequence.