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The work presented in this thesis centres on the development of a work-flow in which coarse-grained molecular dynamics (MD) simulations of a planar phospholipid bilayer, containing membrane proteins, is used to parameterize a larger-scale simplified bilayer model. Using this work-flow, repeat simulations and simulations of larger systems are possible, better enabling the calculation of bulk statistics for the system. The larger-scale simulations can be run on commercial hardware, once the initial parameterization has been performed. In the simplified representation, each protein was initially only represented by the position of its centre of mass and later with the inclusion of its orientation. The membrane protein used throughout most of this work was the bacterial outer membrane protein NanC, a member of the KdgM family of proteins. To parameterize the motion and interaction of proteins using MD, the potential of mean force (PMF) for the pairwise association of two proteins in a bilayer was calculated for a variety of orientational combinations, using a modified umbrella sampling procedure. The relative orientations chosen represented extreme examples of the contact regimes between the two proteins: they approximately corresponded to maxima and minima of the solvent inaccessible surface area, calculated when the proteins were in contact. These PMFs showed that there was a correlation between the buried surface area and the depth of the potential well in the PMF; this is something that, to date, has only been observed in these relatively-'featureless' membrane proteins (but is seen in globular proteins), where the effect of the interactions with lipids in the bilayer plays a larger role. Features in the PMF were observed that resulted from the preferential organization of lipids in the region between the two proteins. These features were small wells in the PMF, which occurred at protein separations that corresponded to the intervening lipids being optimally packed between the proteins. This result further highlighted the role that the lipids in the bilayer played in the interaction between the NanC proteins. The simplified bilayer model was parameterized using the PMFs and the relationship between buried surface area and potential well depth. The initial model included only the proteins' positions. A series of Monte Carlo simulations were performed in order to compare the system behaviour to that of an equivalent MD simulation. Initially, the MD simulation and our parameterized model did not show a good agreement, so a Monte Carlo scheme that incorporated cluster-based movements was implemented. The agreement between the MD simulation and the simulations of our model using the cluster-based scheme, when comparing diffusive and clustering behaviour, was good. Including the orientation-dependent features of the parameterization resulted in the emergence of behaviour that was not clearly detectable in the MD simulation. Finally, attempts were made to parameterize the model using PMFs for the association of rhodopsin from the literature. Rhodopsin was a much more complicated protein to represent: there was not a clear correlation between surface area and the features of the PMF, and the geometry of the interaction between two rhodopsins was more complicated. Simulations of the 'rows-of-dimers' system of rhodopsin, observed in disc membranes, was not entirely well represented by the model; for such a closely packed system, where the number of lipids is much closer to the number of proteins, the use of an implicit-lipid model meant that the effect of the reduced lipid mobility was not adequately captured. However, the model accurately captures the orientational composition of the system. Future work should be focussed on incorporating explicit representations of the lipid in the system so that the behaviour of close-packed systems are better represented.

Gas-expanded liquids (GXLs) are novel and environmentally benign solvent systems with applications in reactions, separations, nanotechnology, drug delivery, and microelectronics. GXLs are liquid mixtures consisting of an organic solvent combined with a benign gas, such as CO2, in the nearcritical regime. In this work, molecular dynamics simulations have been combined with experimental techniques to elucidate the cybotactic region or local environment in gas-expanded liquids. Molecular dynamics simulations show clustering of methanol molecules in carbon dioxide-methanol mixtures. This clustering was not observed in carbon dioxide-acetone mixtures. Furthermore, addition of carbon dioxide enhances diffusivity of solutes in gas-expanded media as shown by both simulations and Taylor-Aris dispersion experiments. Finally, local structure and local compositions around pyrene in carbon dioxide-methanol and carbon-dioxide acetone were investigated using simulations and UV-vis spectroscopy.

The objective of this research is to use atomistic simulations to investigate dislocation nucleation from grain boundaries in face-centered cubic aluminum and copper. This research primarily focuses on asymmetric tilt grain boundaries and has three main components. First, this research uses molecular statics simulations of the structure and energy of these faceted, dissociated grain boundary structures to show that Σ3 asymmetric boundaries can be decomposed into the structural units of the Σ3 symmetric tilt grain boundaries, i.e., the coherent and incoherent twin boundaries. Moreover, the energy for all Σ3 asymmetric boundaries is predicted with only the energies of the Σ3 symmetric boundaries and the inclination angle. Understanding the structure of these boundaries provides insight into dislocation nucleation from these boundaries. Further work into the structure and energy of other low order Σ asymmetric boundaries and the spatial distribution of free volume within the grain boundaries also provides insight into dislocation nucleation mechanisms. Second, this research uses molecular dynamics deformation simulations with uniaxial tension applied perpendicular to these boundaries to show that the dislocation nucleation mechanisms in asymmetric boundaries are highly dependent on the faceted, dissociated structure. Grain boundary dislocation sources can act as perfect sources/sinks for dislocations or may violate this premise by increasing the dislocation content of the boundary during nucleation. Furthermore, simulations under uniaxial tension and uniaxial compression show that nucleation of the second partial dislocation in copper exhibits tension-compression asymmetry. Third, this research explores the development of models that incorporate the resolved stress components on the slip system of dislocation nucleation to predict the atomic stress required for dislocation nucleation from single crystals and grain boundaries. Single crystal simulations of homogeneous dislocation nucleation help define the role of lattice orientation on the nucleation stress for grain boundaries. The resolved stress normal to the slip plane on which the dislocation nucleates plays an integral role in the dislocation nucleation stress and related mechanisms. In summary, the synthesis of various aspects of this work has provided improved understanding of how the grain boundary character influences dislocation nucleation in bicrystals, with possible implications for nanocrystalline materials.

Proteins and DNA are essential to life as we know it and understanding their function is understanding their structure and dynamics. The importance of the latter is being appreciated more in recent years and has led to the development of novel interdisciplinary techniques and approaches to studying protein function. Three techniques to study protein structure and dynamics have been used and combined in different ways in the context of this thesis and have led to a better understanding of the three systems described herein. X-ray crystallography is the oldest and still arguably most popular technique to study macromolecular structures. Nuclear magnetic resonance (NMR) spectroscopy is a not much younger technique that is a powerful tool not only to probe molecular structure but also dynamics. The last technique described herein are molecular dynamics (MD) simulations, which are only just growing out of their infancy. MD simulations are computer simulations of macromolecules based on structures solved by X-ray crystallography or NMR spectroscopy, that can give mechanistic insight into dynamic processes of macromolecules whose amplitudes can be estimated by the former two techniques. MD simulations of the model protein GB3 (B3 immunoglobulin-binding domain of streptococcal protein G) were conducted to identify origins of discrepancies between order parameters derived from different sets of MD simulations and NMR relaxation experiments.The results highlight the importance of time scales as well as sampling when comparing MD simulations to NMR experiments. Discrepancies are seen for unstructured regions like loops and termini and often correspond to nanosecond time scale transitions between conformational substates that are either over- or undersampled in simulation. Sampling biases can be somewhat remedied by running longer (microsecond time scale) simulations. However, some discrepancies persist over even very long trajectories. We show that these discrepancies can be due to the choice of the starting structure and more specifically even differences in protonation procedures. A test for convergence on the nanosecond time scale is shown to be able to correct for many of the observed discrepancies. Next, MD simulations were used to predict in vitro thermostability of members of the bacterial Ribonuclease HI (RNase H) family of endonucleases. Thermodynamic stability is a central requirement for protein function and a goal of protein engineering is improvement of stability, particularly for applications in biotechnology. The temperature dependence of the generalized order parameter, S, for four RNase H homologs, from psychrotrophic, mesophilic and thermophilic organisms, is highly correlated with experimentally determined melting temperatures and with calculated free energies of folding at the midpoint temperature of the simulations. This study provides an approach for in silico mutational screens to improve thermostability of biologically and industrially relevant enzymes. Lastly, we used a combination of X-ray crystallography, NMR spectroscopy and MD simulations to study specificity of the interaction between Drosophila Hox proteins and their DNA target sites. Hox proteins are transcription factors specifying segment identity during embryogenesis of bilaterian animals. The DNA binding homeodomains have been shown to confer specificity to the different Hox paralogs, while being very similar in sequence and structure. Our results underline earlier findings about the importance of the N-terminal arm and linker region of Hox homeodomains, the cofactor Exd, as well as DNA shape, for specificity. A comparison of predicted DNA shapes based on sequence alone with the shapes observed for different DNA target sequences in four crystal structures when in complex with the Drosophila Hox protein AbdB and the cofactor Exd, shows that a combined ”induced fit”/”conformational selection” mechanism is the most likely mechanism by which Hox homeodomains recognize DNA shape and achieve specificity. The minor groove widths for all sequences is close to identical for all ternary complexes found in the different crystal structures, whereas predicted shapes vary between the different DNA sequences. The sequences that have shown higher affinity to AbdB in vitro have a predicted DNA shape that matches the observed DNA shape in the ternary complexes more closely than the sequences that show low in vitro affinity to AbdB. This strongly suggests that the AbdB-Exd complex selects DNA sequences with a higher propensity to adopt the final shape in their unbound form, leading to higher affinity. An additional AbdB monomer binding site with a strongly preformed binding competent shape is observed for one of the oligomers in the reverse complement strand of one of the canonical (weak) Hox-Exd complex binding site. The shape preference seems strong enough for AbdB monomer binding to compete with AbdB-Exd dimer binding to that same oligomer, suggested by the presence of both binding modes in the same crystal. The monomer binding site is essentially able to compete with the dimer binding site, even though binding with the cofactor is not possible, because its shape is very close to the ideal shape. A comparison of different crystal structures solved herein and in the literature as well as a set of molecular dynamics simulations was performed and led to insights about the importance of residues in the Hox N-terminal arm for the preference of certain Hox paralogs to certain DNA shapes. Taken together all these insights contribute to our understanding of Hox specificity in particular as well as protein-DNA interactions in general.

Yan, Changqing. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 92-97). / Abstracts in English and Chinese. / ABSTRACT (ENGLISH) --- p.iii / ABSTRACT (CHINESE) --- p.iv / ACKNOWLEDGMENTS --- p.v / LIST OF ABBREVIATIONS --- p.vi / TABLE OF CONTENTS --- p.vii / LIST OF FIGURES --- p.ix / LIST OF TABLES --- p.x / Chapter CHAPTER ONE: --- BACKGROUND INFORMATION --- p.1 / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.2 --- Deoxyribonucleic Acid --- p.2 / Chapter 1.3 --- DNA Studies --- p.9 / Chapter 1.4 --- Cisplatin Studies --- p.11 / Chapter 1.5 --- Scope of the Thesis --- p.13 / Chapter CHAPTER TWO: --- METHODOLOY AND COMPUTATION --- p.16 / Chapter 2.1 --- Introduction --- p.16 / Chapter 2.2 --- Molecular Dynamics Simulation --- p.16 / Chapter 2.3 --- Quantum Mechanics Calculation --- p.23 / Chapter 2.4 --- Verification of Methodology --- p.25 / Chapter 2.4.1 --- Backbone Torsion Angles --- p.25 / Chapter 2.4.2 --- N7-N7 Distance --- p.30 / Chapter 2.4.3 --- Location of HOMO --- p.33 / Chapter 2.5 --- Summary --- p.35 / Chapter CHAPTER THREE: --- UNDERSTANDING OF THE CISPLATIN-DNA CROSSLINKS --- p.36 / Chapter 3.1 --- Introduction --- p.36 / Chapter 3.2 --- MO Analysis --- p.37 / Chapter 3.3 --- Potential Binding Products with the Ligand --- p.37 / Chapter 3.3.1 --- "1,2-d(GpG) Intrastrand Crosslink" --- p.43 / Chapter 3.3.2 --- "l,2-d(ApG) Intrastrand Crosslink" --- p.43 / Chapter 3.3.3 --- "l,3-d(GpXpG) Intrastrand Crosslink" --- p.44 / Chapter 3.3.4 --- d(GpC)d(GpC) Interstrand Crosslink --- p.44 / Chapter 3.3.5 --- d(GpXpC)d(GpXpC) Interstrand Crosslink --- p.44 / Chapter 3.3.6 --- Summary --- p.45 / Chapter 3.4 --- Potential Binding Products Analysis --- p.47 / Chapter 3.4.1 --- Site Identification Convention --- p.47 / Chapter 3.4.2 --- Potential Binding Products Analysis --- p.48 / Chapter 3.4.3 --- Applications --- p.53 / Chapter 3.5 --- Cisplatin-DNA Crosslink Products Analysis --- p.56 / Chapter 3.5.1 --- "1,2-d(GpG) and l,2-d(ApG) Intrastrand Crosslinks" --- p.61 / Chapter 3.5.2 --- "l,3-d(GpXpG) Intrastrand and d(GpXpC)d(GpXpC) Interstrand Crosslinks" --- p.62 / Chapter 3.5.3 --- d(GpC)d(GpC) Interstrand Crosslinks --- p.63 / Chapter 3.5.4 --- Platination at Terminal Positions --- p.65 / Chapter 3.6 --- Summary --- p.65 / Chapter CAHPTER FOUR: --- CONCLUDING REMARKS --- p.67 / APPENDIX I: BACKBONE TORSION ANGLES AND SUGAR RING CONFORMATIONS OF THE OPTIMIZED GEOMETRIES --- p.69 / APPENDIX II: BACKBONE TORSION ANGLES OF THE EXPERIMENTAL SEQUENCES FROM NUCLEIC ACID DATABASE (NDB) --- p.77 / REFERENCES --- p.92

The mechanical and physical properties of polymers are determined primarily by the underlying nano-scale structures and characteristics such as entanglements, crystallites, and molecular orientation. These structures evolve in complex manners during the processing of polymers into useful articles. Limitations of available and foreseeable computational capabilities prevent the direct determination of macroscopic properties directly from atomistic computations. As a result, computational tools and methods to bridge the length and time scale gaps between atomistic and continuum models are required. In this research, an internal state variable continuum model has been developed whose internal state variables (ISVs) and evolution equations are related to the nano-scale structures. Specifically, the ISVs represent entanglement number density, crystal number density, percent crystallinity, and crystalline and amorphous orientation distributions. Atomistic models and methods have been developed to investigate these structures, particularly the evolution of entanglements during thermo-mechanical deformations. A new method has been created to generate atomistic initial conformations of the polymer systems to be studied. The use of the hyperdynamics method to accelerate molecular dynamics simulations was found to not be able to investigate processes orders of magnitude slower that are typically measurable with traditional molecular dynamics simulations of polymer systems. Molecular dynamics simulations were performed on these polymer systems to determine the evolution of entanglements during uniaxial deformation at various strain rates, temperatures, and molecular weights. Two methods were evaluated. In the first method, the forces between bonded atoms along the backbone are used to qualitatively determine entanglement density. The second method utilizes rubber elasticity theory to quantitatively determine entanglement evolution. The results of the second method are used to gain a clearer understanding of the mechanisms involved to enhance the physical basis of the evolution equations in the continuum model and to derive the models material parameters. The end result is a continuum model that incorporates the atomistic structure and behavior of the polymer and accurately represents experimental evidence of mechanical behavior and the evolution of crystallinity and orientation.

Despite numerous advances in experimental methodologies capable of addressing the various phenomenon occurring on metal surfaces, atomic scale resolution of the microscopic dynamics remains elusive for most systems. Computational models of the processes may serve as an alternative tool to fill this void. To this end, parallel molecular dynamics simulations of self-diffusion on metal surfaces have been developed and employed to address microscopic details of the system. However these simulations are not without their limitations and prove to be computationally impractical for a variety of chemically relevant systems, particularly for diffusive events occurring in the low temperature regime. To circumvent this difficulty, a corresponding coarse-grained representation of the surface is also developed resulting in a reduction of the required computational effort by several orders of magnitude, and this description becomes all the more advantageous with increasing system size and complexity. This representation provides a convenient framework to address fundamental aspects of diffusion in nonequilibrium environments and an interesting mechanism for directing diffusive motion along the surface is explored. In the ensuing discussion, additional topics including transition state theory in noisy systems and the construction of a checking function for protein structure validation are outlined. For decades the former has served as a cornerstone for estimates of chemical reaction rates. However, in complex environments transition state theory most always provides only an upper bound for the true rate. An alternative approach is described that may alleviate some of the difficulties associated with this problem. Finally, one of the grand challenges facing the computational sciences is to develop methods capable of reconstructing protein structure based solely on readily-available sequence information. Herein a checking function is developed that may prove useful for addressing whether a particular proposed structure is a viable possibility.

Molecular dynamics (MD) simulations were used to investigate the structural basis for the functions of three proteins: Fc(gamma) receptor III (CD16), von Willebrand factor (VWF), and integrin. CD16, a heavily glycosylated protein expressed on human immune cells, plays a crucial role in immune defense by linking antibody-antigen complexes with cellular effector functions. Glycosylation of CD16 decreases its affinity for IgG. MD simulations were run for CD16-IgG Fc complexes with or without an N-glycan on CD16. The two simulated complexes show different conformations. Molecular Mechanics-Poisson Boltzmann Surface Area (MM-PBSA) approach was used to calculate the binding free energy of the CD16-IgG Fc complexes. The calculated binding free energy helped to identify critical residues. VWF, a multimeric multidomain glycoprotein, initiates platelet adhesion at the sites of vascular injury. A specific VWF metalloprotease, A Disintegrin And Metalloprotease with ThromboSpondin motifs member 13 (ADAMTS-13), cleaves the Tyr1605-Met1606 bond in the VWF A2 domain to generate the full spectrum of plasma VWF species. Shear stress or denaturants assist VWF cleavage by ADAMTS-13 due to the unfolding of A2. MD was used to simulate the unfolding processes of A2 under force or high temperature. The beta-strands of A2 were pulled out sequentially by force, during which the cleavage site changed in steps from the fully buried state to the fully exposed state. Thermal unfolding follows a very different pathway. Integrins are adhesion molecules mediating cell-cell, cell-extracellular matrix, and cell-pathogen interactions. Experiments suggest that integrins can undergo a large-scale change from a bent to an extended conformation, associating with a transition from low to high affinity states, i.e., integrin activation. Steered MD was utilized to simulate the bent-to-extended conformational transition in time of aVb3 integrin. The integrin was observed to change smoothly from the bent to the extended conformation. One major energy barrier was overcome, corresponding to the disruption of the interactions at Hybrid/EGF4/bTD interfaces. A partially extended conformation tends to bend back while a fully extended conformation is stabilized by the coordination of Asp457 with Ca2+ at alpha-genu. Unbending with separated legs overcomes more energy barriers.