The Archaeal MCM Helicase

DNA复制是生物体繁殖和进化所必需的。DNA复制机制确保了细胞分裂过程中遗传信息的复制和传递。此外，该过程需要精确，并且每个细胞周期必须发生一次且仅发生一次。这个过程在所有生命形式中在功能上和结构上都是保守的，可以分为三个阶段：起始，延伸和终止。DNA复制已经在细菌、病毒和噬菌体中得到了很好的表征，而真核生物和古细菌中的许多方面仍然是未知的。特别是，结构和生物化学性质的蛋白质和酶参与的起始过程仍有待描述。本研究将集中在参与起始和延伸的关键酶--微小染色体维持（MCM）解旋酶。将采用各种技术来确定MCM解旋酶的结构和生物化学性质的古菌甲烷嗜热菌thermautotrophicus。这些研究将有助于理解古细菌中的DNA复制。此外，由于古细菌MCM解旋酶的生化性质与真核生物解旋酶相似，该研究也将有助于了解真核生物中的复制。在这项研究中所产生的结果与MCM解旋酶从其他古菌的比较将提供更丰富的理解的复制机制的演变。具体来说，研究将集中在1）MCM活性形式和结构的测定，2）酶的生化性质的阐明，3）与解旋酶相互作用的蛋白质的鉴定和分离，以及它们对解旋酶活性的影响。该项目在招收各级学生并为他们提供真正的研究经验方面有着悠久而杰出的历史。目前的项目将为两名研究生、四名高中生（每个项目年一名）和八名本科生（每个项目年两名）提供培训。还有直接接触到蒙哥马利学院的生物技术计划，这是马里兰州最大和最多样化的社区学院。

DNA replication is required for the propagation and evolution of living organisms. The mechanism of DNA replication ensures the duplication and transfer of genetic information during cell division. Furthermore, the process needs to be precise, and must occur once and only once per cell cycle. The process is functionally and often structurally conserved in all life forms and can be divided into three phases: initiation, elongation, and termination. DNA replication has been well characterized in bacteria, viruses and bacteriophages, whereas many aspects of the process in eukarya and archaea are still unknown. In particular, the structures and biochemical properties of the proteins and enzymes involved in the initiation process remain to be described. This research will focus on a key enzyme participating in initiation and elongation -- the minichromosome maintenance (MCM) helicase. Various techniques will be employed to determine the structure and biochemical properties of the the MCM helicase from the archaeon Methanothermobacter thermautotrophicus. These studies will contribute significantly to the understanding of DNA replication in archaea. In addition, because the biochemical properties of the archaeal MCM helicase are similar to those of the eukaryotic helicase, the study will also contribute to the understanding of replication in eukarya. Comparison of the results generated in this study with those obtained with MCM helicases from other archaea will provide a richer understanding of the evolution of the replication machinery. Specifically, the study will focus on 1) the determination of the MCM active form and structure, 2) the elucidation of the biochemical properties of the enzyme, and 3) the identification and isolation of proteins that interact with the helicase, and their effect on helicase activity.Although CARB is a research institution without easy access to students, this project has a long and distinguished history of recruiting students at all levels and providing them with a genuine research experience. The current project will provide training for two graduate students, four high school students (one per project year) and eight undergraduate students (two per project year). There is also direct out reach to the biotechnology program at Montgomery College, the largest and most diverse community college in the Maryland.