SV40 large T antigen (LT) is a potent carcinogen, and plays an essential role for SV40 DNA replication as the replicative helicase and replication initiator protein. SV40 replication serves as a model for eukaryotic DNA replication, as SV40 uses all the essential cellular replication proteins (primase, polymerase, PCNA, Topoisomerases, etc.), except for the helicase and cellular initiator proteins that consist of multiple initiator factors (such as Orc, Ctd1, Cdc6, MCM in eukaryotes, or DnaA/DnaC/DnaB in prokaryotes) to initiate DNA replication, i.e. marking the replication origin, recruiting helicase, melting origin, and activating helicase. For SV40 replication, LT alone fulfills essentially all the initiator functions and is the helicase for replication fork unwinding during elongation phase. The long-term goal of this research is to understand how LT functions as a helicase to coordinate the functions of the other replication proteins for DNA replication, as well as how LT transforms cells.
Specific aims are designed to understand how LT hexameric and double hexameric helicase melt the origin DNA and unwinds dsDNA to initiate DNA replication. We plan to use mainly X-ray protein crystallography, assisted with EM and AFM, single molecule assay, computational method, molecular biology and functional biochemistry in vitro and in cells. The results from this research are expected to have potential impact on the field of DNA replication in eukaryotic cells and on cancer biology.
SV40 large T (LT) has remarkably diverse biological activities. Besides its ability to regulate many aspects of viral infection and cellular processes, LT in its hexameric and double hexameric forms is an efficient molecular machine that can melt dsDNA and unwind replication forks for DNA replication. LT is the only viral protein required for SV40 minichromosome DNA replication, all the rest proteins are from cellular replication machinery in mammalian cells. In this minichromosome replication, LT performs the functions of the cellular helicase MCM and several other initiator proteins for origin localization and melting. As a result, SV40 replication system has been serving as a model system for studying eukaryotic replication. The study of LT helicase mechanisms will have general implications for understanding other replicative helicases, especially for those from eukaryotic cells. We aim to understand the detailed molecular mechanisms of the helicase function of LT hexameric and double hexameric machine. The data generated from this research will provide valuable information about helicase function and DNA replication. This study bears high relevance to cancer biology.
|Wang, Damian; Álvarez-Cabrera, Ana Lucia; Chen, Xiaojiang S (2017) Study of SV40 large T antigen nucleotide specificity for DNA unwinding. Virol J 14:79|
|Astumian, R Dean; Mukherjee, Shayantani; Warshel, Arieh (2016) The Physics and Physical Chemistry of Molecular Machines. Chemphyschem 17:1719-41|
|Gai, Dahai; Wang, Damian; Li, Shu-Xing et al. (2016) The structure of SV40 large T hexameric helicase in complex with AT-rich origin DNA. Elife 5:|
|Xia, Yisui; Niu, Yanling; Cui, Jiamin et al. (2015) The Helicase Activity of Hyperthermophilic Archaeal MCM is Enhanced at High Temperatures by Lysine Methylation. Front Microbiol 6:1247|
|Mukherjee, Shayantani; Bora, Ram Prasad; Warshel, Arieh (2015) Torque, chemistry and efficiency in molecular motors: a study of the rotary-chemical coupling in F1-ATPase. Q Rev Biophys 48:395-403|
|Mukherjee, Shayantani; Warshel, Arieh (2015) Dissecting the role of the ?-subunit in the rotary-chemical coupling and torque generation of F1-ATPase. Proc Natl Acad Sci U S A 112:2746-51|
|Mukherjee, Shayantani; Warshel, Arieh (2015) Brønsted slopes based on single-molecule imaging data help to unveil the chemically coupled rotation in F1-ATPase. Proc Natl Acad Sci U S A 112:14121-2|
|Plotnikov, Nikolay V; Prasad, B Ram; Chakrabarty, Suman et al. (2013) Quantifying the mechanism of phosphate monoester hydrolysis in aqueous solution by evaluating the relevant ab initio QM/MM free-energy surfaces. J Phys Chem B 117:12807-19|
|B, Ram Prasad; Plotnikov, Nikolay V; Lameira, Jeronimo et al. (2013) Quantitative exploration of the molecular origin of the activation of GTPase. Proc Natl Acad Sci U S A 110:20509-14|
|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|
Showing the most recent 10 out of 29 publications