Structural Studies of DNA Replication Proteins by X-Ray Crystallography and Molecular Dynamics

Organisms that span evolutionary space utilize an assembly of proteins (the replicase) in a coordinated effort to faithfully replicate their genomes. This chromosomal replicase consists of three functionally distinct subassemblies. The first of these is the polymerase/exonuclease complex, which harbors DNA synthesis and proofreading activities. The second functional complex is the sliding clamp which adopts a ring-shaped structure, composed of either two or three protomers. It confers processivity onto the polymerase subunit by tethering it to the template. The third complex is the clamp loader complex, which loads the sliding clamp onto DNA using energy from ATP binding and hydrolysis. This thesis analyses the structures of two important components of the chromosomal replicase assembly through X-ray crystallography and molecular dynamics simulations. First, a crystal structure of the DNA polymerase from archaebacterium Desulfurococcus Tok (D. Tok Pol) was solved at 2.4 A. The structure revealed its similarity to that of the DNA polymerase from bacteriophage RB69 in spite of a low sequence identity between these two members of the Pol II family of DNA polymerases. Secondly, a series of molecular dynamics simulations were performed on the sliding clamps from Escherichia coli and Streptococcus pyogenes. The studies demonstrated that one subunit of the dimeric clamp, when the other subunit is absent, would relax to a structure of reduced curvature ( open state) when compared to its structure in the dimer ( closed state). Free energy calculations suggest that this spontaneous structural change is driven by higher angle and dihedral energies in the closed state. This finding led to the hypothesis that sliding clamps are spring-loaded rings that relax during the loading reaction when one of their oligomeric interfaces is disrupted by the clamp loader complex. Lastly, deconvolution of X-ray diffraction data from a perfectly merohedrally twinned crystal was used to improve the structure of the human sliding clamp in complex with a peptide derived from p2iWAF1/clp\ a DNA replication inhibitor, from a resolution of 2.6 A to a resolution of 2.3 A

Asymptotically Compatible Schemes for Nonlocal Ohta Kawasaki Model

We study the asymptotical compatibility of the Fourier spectral method in multidimensional space for the Nonlocal Ohta-Kawasaka (NOK) model, which is proposed in our previous work. By introducing the Fourier collocation discretization for the spatial variable, we show that the asymptotical compatibility holds in 2D and 3D over a periodic domain. For the temporal discretization, we adopt the second-order backward differentiation formula (BDF) method. We prove that for certain nonlocal kernels, the proposed time discretization schemes inherit the energy dissipation law. In the numerical experiments, we verify the asymptotical compatibility, the second-order temporal convergence rate, and the energy stability of the proposed schemes. More importantly, we discover a novel square lattice pattern when certain nonlocal kernel are applied in the model. In addition, our numerical experiments confirm the existence of an upper bound for the optimal number of bubbles in 2D for some specific nonlocal kernels. Finally, we numerically explore the promotion/demotion effect induced by the nonlocal horizon, which is consistent with the theoretical studies presented in our earlier work