7-deazaguanines in DNA: mechanism and structure of complex genome modification

总结。酶导向的核苷酸修饰是核酸的一个关键成分 所有生物体都使用它来满足多种基本需求 信息传输系统，包括DNA保护、翻译保真度、RNA稳定、 和表观遗传调控。在RNA和DNA之间的串扰的一个显著例子中 处理过程中，我们最近发现已知的最复杂的修饰系统之一 发生在RNA中，负责7-去氮鸟嘌呤修饰队列(Q)和 考古辛(G+)也被各种生物用来修饰DNA。事实上，在 细菌由大约10种蛋白质组成的一个精细的限制-修饰(Rm)系统。 基于2‘-脱氧-7-氰基和2’-脱氧-7-氨基-7-脱氮鸟苷(DPreQ0)的形成 和dADG)，以及这些核苷，以及dG+和2‘-脱氧-7- 氨甲基-7-脱氮鸟苷(DPreQ1)，也在噬菌体和 古生物病毒。在确定了参与形成7- 基于去氮鸟嘌呤的DNA修饰，我们现在提议阐明其分子基础 因为他们的功能。具体目标1和2中描述的研究涉及修改 细菌中的一种蛋白质DpdA、DpdB和DpdC，它们共同负责 DPreQ0和dADG的形成，并作为7-去氮鸟嘌呤修饰的模型 在DNA中。在具体目标3中，我们将更广泛地了解DPDA系列，并考虑以下系统 缺乏Dpdb，包含DpdAC融合，以及噬菌体DpdA，以便更好地理解 细菌修饰中需要DpdB的潜在ATPase活性及其结构基础 在细菌和病毒系统中具有序列特异性。

SUMMARY. Enzymatically-directed nucleotide modification is a key component of nucleic acid processing and is used by all organisms to address a multitude of fundamental needs in information transfer systems, including protection of DNA, translational fidelity, RNA stabilization, and epigenetic regulation. In a remarkable example of the cross-talk between RNA and DNA processing, we recently discovered that one of the most complex modification systems known to occur in RNA, that responsible for the 7-deazaguanine modifications queuosine (Q) and archaeosine (G+), is also utilized by diverse organisms for the modification of DNA. Indeed, in Bacteria a set of roughly 10 proteins comprise an elaborate restriction-modification (RM) system based on the formation of 2’-deoxy-7-cyano- and 2’-deoxy-7-amido-7-deazaguanosine (dPreQ0 and dADG, respectively) in DNA, and these nucleosides, as well as dG+ and 2’-deoxy-7- aminomethyl-7-deazaguanosine (dPreQ1), have also been found in the DNA of phage and archaeal viruses. Having identified the relevant proteins involved in the formation of 7- deazaguanine based DNA modifications, we are now proposing to elucidate the molecular basis for their function. The research described in specific aims 1 & 2 address the modification machinery in bacteria, the proteins DpdA, DpdB, and DpdC, which together are responsible for the formation of dPreQ0 and dADG, and serve as a model for 7-deazaguanine based modification in DNA. In specific aim 3 we take a broader look at the DpdA family and consider systems that lack DpdB and contain DpdAC fusions, as well as a phage DpdA in order to better understand the need for the cryptic ATPase activity of DpdB in bacterial modification, and the structural basis of sequence specificity in the bacterial and viral systems.