Structural Studies of MCM Complex

描述（由申请人提供）：微染色体维持蛋白（MCM）复合物作为解旋酶发挥作用，解绕DNA为DNA复制提供模板，并在调节细胞生长中起关键作用。MCM解旋酶被认为是细胞DNA复制的许可因子。真核生物和古细菌的MCM蛋白形成六聚体环状复合物，MWt约为0.5-1.0兆多尔顿。尽管付出了广泛的努力，但我们对MCM如何解绕DNA， MCM在解绕过程中如何与DNA相互作用，以及ATP如何利用来驱动解绕所需的MCM构象变化的理解存在空白。MCM在古细菌和真核生物中保守。真核MCM包含六个同源物，形成异六聚体和可能的高阶复合物。然而，在一些古生菌中，MCM复合体只含有一种MCM蛋白，可以组装成同型六聚体或十二聚体。因此，古细菌MCM同源低聚物为理解MCM复合物的结构/功能提供了一个更简单的系统。本研究的目标是利用古细菌MCM复合物作为模型系统来了解MCM复合物的结构/功能，特别强调MCM蛋白的寡聚化、atp触发的构象变化以及DNA结合和重塑功能的机制。结构生物学，生物物理学和功能生物化学的结合将被用于研究。所得到的数据将为理解DNA在复制叉上的解绕机制提供有价值的信息。它还将为同源真核MCM复合体提供结构/功能方面的见解，这与细胞生长调节和癌症生物学具有高度相关性。公共卫生相关性：所有生物体的一个基本过程是复制基因组DNA（或DNA复制），以确保正确的遗传信息传递。为了复制基因组DNA，需要解旋酶来解压缩双螺旋DNA (dsDNA)，以便在复制过程中为子链合成提供ssDNA模板。MCM复合体是一种解旋酶，它可以解绕dsDNA，对DNA复制和细胞生长至关重要。我们建议结合结构生物学、生物物理学和功能生物化学等方法来研究这种重要的MCM解旋酶的结构/功能，以了解这种酶如何利用ATP的能量来解开双螺旋DNA进行复制。

DESCRIPTION (provided by applicant): Minichromosome maintenance protein (MCM) complex functions as a helicase that unwinds DNA to provide template for DNA replication and plays a critical role in regulating cell growth. MCM helicases are considered to be a licensing factor for cellular DNA replication. MCM proteins from eukaryotes and archaea form hexamers ring complex, with MWt approximating 0.5-1.0 Mega Dolton. Despite extensive efforts, gap of knowledge exists in our understanding of how MCM unwinds DNA, how MCM interacts with DNA during unwinding, and how ATP is utilized to drive the conformational changes of MCM needed for unwinding. MCM are conserved in archaea and eukaryotes. Eukaryotic MCM contains six homologs that form hetero-hexamer and possibly higher order complexes. However, the MCM complex in some archaea contains only one MCM protein that can assemble into homo-hexamers or dodecamers. Thus, archaeal MCM homo-oligomers provide a simpler system for understanding the structure/function of MCM complex. The goal of this proposal is to use archaeal MCM complexes as a model system to understand the structure/functions of MCM complex, with particular emphasis on the mechanisms regarding oligomerization, ATP-triggered conformational changes, and DNA binding and remodeling functions of MCM proteins. A combination of structural biology, biophysics, and functional biochemistry will be employed for the study. The resulting data will provide valuable information for understanding the DNA unwinding mechanism at the replication fork. It will also provide structural/functional insights for the homologous eukaryotic MCM complex, which bears high relevance to cell growth regulation and cancer biology. PUBLIC HEALTH RELEVANCE: One essential process for all living organisms is duplicate the genomic DNA (or DNA replication) to ensure correct genetic information passage. To duplicate genomic DNA, helicases is required to unzip the double helix DNA (dsDNA) to provide ssDNA template for daughter strand synthesis in the replication process. MCM complex is such a helicase that unwinds dsDNA and is critical for DNA replication and cell growth. We propose to study the structure/function of this important MCM helicase using a combination of approaches including structural biology, biophysics, and functional biochemistry, in order to understand how this enzyme utilizes the energy of ATP to unwind the double helix DNA for replication.