Simulating the helicase motor of SV40 large tumor antigen

Helicases are motor protein that utilize the energy derived from NTP binding and hydrolysis to translocate and unwind DNA/RNA during the replication. Understanding the energy coupling of NTP hydrolysis cycle to the DNA movement is the key to understand the DNA replication mechanism in the molecular motor. The helicase domain of simian virus 40 large tumor antigen (SV40 LTag) is a ring-shaped AAA+ domain that participates in viral DNA replication and host cell growth control. Recent SV40 LTag structure studies have provided a set of high resolution structures in different nucleotide binding states. Hence, in this thesis we use LTag helicase as a model protein, and present the first systematic simulation study on the mechanism of the LTag helicase motor. Our work includes three major sections: first, we model the LTag ATPase activity and the helicase activity based on the biochemistry experiment results. This model indicates that the LTag helicase subunits work in highly cooperative patterns. When the origin DNA is presented, the helicase translocates DNA in a sequential pattern. When the fork DNA is added, the helicase works in a semi-sequential pattern, otherwise, the subunit cooperativity is not significant. Second, we present the first simulation study on the ATP binding/hydrolsis procedure using the non-equilibrated molecular dynamics method, the results suggest a three-stage Locker-binding model. We evaluate the energy profile using the LRA version of the semi-microscopic Protein Dipoles-Langvin Dipoles method (PDLD/S). The energy profile matches the experimental results. Thirdly, we investigate the electrostatic energy that guides the single-strand DNA (ssDNA) translocation process and propose a unidirectional translocation model. To accomplish this work, an ssDNA/LTag complex model is built using the structure information from the LTag helicase and the E1 protein-DNA complex, a two-dimensional effective electrostatic free-energy landscape is calculated based on the ssDNA/LTag model, and the unidirectional model is proposed by evaluating the energy landscape. The time dependence of the coupled protein-DNA motion is explored by simulating the translocation process using a renormalized method. Altogether, our theoretical and simulation study advanced our understanding of the fundamental molecular mechanism underlying the directional movement of ring-shaped helicase motor.