Structural studies of MCM complex 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

Title: Structural studies of MCM complex 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.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM080338-04
Application #
8204543
Study Section
Macromolecular Structure and Function C Study Section (MSFC)
Program Officer
Preusch, Peter C
Project Start
2009-01-01
Project End
2013-11-30
Budget Start
2011-12-01
Budget End
2013-11-30
Support Year
4
Fiscal Year
2012
Total Cost
$325,492
Indirect Cost
$124,571
Name
University of Southern California
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
072933393
City
Los Angeles
State
CA
Country
United States
Zip Code
90089
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Xu, Meng; Chang, Y Paul; Chen, Xiaojiang S (2013) Expression, purification and biochemical characterization of Schizosaccharomyces pombe Mcm4, 6 and 7. BMC Biochem 14:5
Brewster, Aaron S; Slaymaker, Ian M; Afif, Samir A et al. (2010) Mutational analysis of an archaeal minichromosome maintenance protein exterior hairpin reveals critical residues for helicase activity and DNA binding. BMC Mol Biol 11:62
Brewster, Aaron S; Chen, Xiaojiang S (2010) Insights into the MCM functional mechanism: lessons learned from the archaeal MCM complex. Crit Rev Biochem Mol Biol 45:243-56