Structural, biochemical and biophysical analysis of DisA and DAC-domain proteins

DNA完整性的维持是维持生命的一个重要因素，细胞通过各种DNA损伤传感器和检查点蛋白来监测这一点。虽然人类的检查点控制非常复杂，而且在机械上了解得很少，但细菌系统有一个简单得多的系统，这是一个很有前途的结构和机械分析的模型系统。我们已经解决了一种新发现的细菌DNA损伤传感器蛋白(DISA)的晶体结构，并发现它与DNA重组中间结构结合，从而阻止细胞进入产孢期。DISA被发现具有以前未被表征的二腺苷环化酶活性。由此产生的c-di-AMP与细菌生理学中众所周知的c-di-GMP有关。C-di-AMP以前没有被描述过，也没有c-di-AMP调节的蛋白已知，这为细菌信号转导开辟了一个全新的领域。最近发现c-di-AMP能在李斯特菌感染时触发细胞的干扰素反应，这可能是由刺蛋白识别的，强调了它在感染生物学和先天免疫中也很重要。我们的目标是分析DISA和其他蛋白质合成c-di-AMP的机制，并确定可能的相互作用蛋白质。作为一个长期目标，我们的目标是识别c-di-AMP信号通路中的蛋白质，以更全面地了解哪些过程可能在细胞中受到调控。

The maintenance of DNA integrity is an important factor in sustainment of life, and cells monitor this by a variety of DNA damage sensors and checkpoint proteins. While humans have a very complex and mechanistically poorly understood checkpoint control, bacterial systems have a much simpler system that is a promising model system for structural and mechanistic analysis. We have solved the crystal-structure of a newly identified bacterial DNA-damage sensor protein (DisA) and discovered that it binds to DNA recombination intermediate structures, thereby preventing the cell from entering sporulation phase. DisA was found to have a previously uncharacterized di-adenylate cyclase enzymatic activity. The resulting c-di-AMP is related to the well know c-di-GMP in bacterial physiology. The fact that c-di-AMP has not been described before and no c-di-AMP-regulated proteins are known, opens up a completely new field in bacterial signaling. C-di-AMP has recently been shown to trigger interferon response of cells upon Listeria infection probably recognized by the STING protein, underlining that it is important also in infection biology and innate immunity. We aim to analyze the mechanism of DisA and other proteins synthesizing c-di-AMP and identify possible interacting proteins. As a long term goal we aim to identify proteins in the c-di-AMP signaling pathway to get a more complete picture of which processes may be regulated in the cell.