Synthetic and Computational Studies on Polycyclic Aromatic Hydrocarbon Derivatives, Nucleoside Analogs and Peptides

In recent years, the understanding of the structure and functions of biological macromolecules has advanced rapidly, the result of which is a better mechanistic understanding of many biological processes. As an outgrowth of this understanding, organic molecules that react with biological macromolecules (DNA) or adopt conformations responsible for specific functions in biological macromolecules (peptides and proteins) have been synthesized and computational modeling studies performed. Polycyclic aromatic hydrocarbons (PAHs) and β-peptides are among synthetic organic compounds known to interact with natural biological macromolecules. This interaction may affect the specific biological functions of the biomacromolecules. A variety of synthetic methodologies have been employed in the synthesis of benzo[c]phenanthrene derivatives, single electron oxidation nucleoside adducts and deoxynucleoside derivatives (Part 1). In Part 2 heterogeneous backbone oligomers containing the β-amino acid, trans-2-aminocyclohexanecarboxylic acid (ACHC), and α-amino acids Ala, Phe, Val, Lys, and Tyr in an alternating sequence have been synthesized. Computational modeling studies have been applied in studying the diastereoselectivity of reaction intermediates in the PAH syntheses (Part 1), the interaction between the organic compounds and biomacromolecules (β-peptides with proteins Fos and Jun, Part 2), and the conformational preference (conformations of α/β-peptides, Part 2). Computational modeling based on molecular and quantum mechanical techniques were applied to complement the syntheses in Parts 1 and 2

Modelling of crystal structure of cis-1,2,3,6 and 3,4,5,6-tetrahydrophthalic anhydrides using lattice energy calculations

Lattice energy calculations using a model potential were performed to model the crystal structures of cis-1,2,3,6- and 3,4,5,6-tetrahydrophthalic (THP) anhydrides. The optimized molecular models using the DFT method at the B3LYP/6-31G** level were found consistent with the available experimental evidence and allowed all differences observed in crystal packing between cis-1,2,3,6- and 3,4,5,6-THP anhydrides to be reproduced. Calculations provide evidence for the presence of dipole–dipole C=O?C=O intermolecular interactions and support the idea that the molecules distort from their ideal geometries, improving packing in both crystals. The search for minima in the lattice energy of both crystals amongst the more common space groups with Z’?=?1, using a simulated annealing crystal structure prediction procedure followed by lattice energy minimization showed that the observed structure of 3,4,5,6-THP anhydride (Z’?=?2) is the thermodynamically most stable, and allowed us to justify why 3,4,5,6-THP anhydride crystallizes in such a complex structure with 16 molecules in the unit cell. The computational model was successful in predicting the second observed form at 173 K for cis-1,2,3,6-THP anhydride as a polymorph, and could predict several hypothetical structures with Z’?=?1 that appear competitive with the observed structures. The results of phonon estimates of zero point intermolecular vibrational energy and entropy suggest that crystal structures of cis-1,2,3,6-THP anhydride cannot be predicted solely on the basis of lattice energy; factors other than thermodynamics favor the observed structures

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

Molecular modelling of thymidylate synthase and rational design of its inhibitors as novel anticancer drugs

In search of novel anticancer drugs, putative inhibitors of the enzyme thymidylate synthase were investigated. The dissertation presents several steps of computationally aided drug design. Two targets are described: active site of the enzyme, for competitive inhibitors, and an allosteric pocket at the dimer interface. The potential hits were selected by computational high-throughput screening (molecular docking calculations) of available drug and prodrug databases. The selected compounds were then modified and scored further to indicate potential leads. Molecular dynamics simulations were performed for selected putative inhibitors of thymidylate synthase, both competitive and allosteric, in order to assess their dynamical behaviour, binding properties and arrangement of the ligands, and to select lead compounds for further tests in vitro. Moreover, a library of peptoids is described, created with the aim to design novel compounds with the desired peptide-like properties. Furthermore, quantum mechanics calculations were conducted to aid the synthesis and investigation of novel enzyme inhibitors, including boron containing compounds.W poszukiwaniu leków przeciwnowotworowych nowej generacji badano potencjalne inhibitory enzymu syntazy tymidylanowej. Opisano szereg etapów komputerowo wspomaganego projektowania leków. Wybrano dwa miejsca docelowe dla poszukiwanych inhibitorów: kieszeń aktywną enzymu oraz kieszeń allosteryczną między podjednostkami białka. Potencjalnie obiecujące związki wybrano w drodze wysokowydajnej procedury przesiewania (przy zastosowaniu metod dokowania molekularnego) dostępnych baz danych leków i proleków, a następnie modyfikację i dalszą selekcję wyników dokowania. Dla wybranych potencjalnych inhibitorów syntazy tymidylanowej, zarówno kompetycyjnych, jak i allosterycznych, przeprowadzono symulacje metodą dynamiki molekularnej w celu oceny dynamiki układu, parametrów wiązania i ułożenia ligandów, jak również wskazania wiodących związków do dalszych badań in vitro. Ponadto opisano bibliotekę peptoidów, stworzoną w celu projektowania nowej generacji związków o pożądanych właściwościach peptydomimetycznych. Wykonano również obliczenia metodami mechaniki kwantowej mające na celu wspomaganie badań i syntezy nowych inhibitorów enzymów, w tym związków zawierających bor

Structural Studies of the Anti-HIV Human Protein APOBEC3G Catalytic Domain: A Dissertation

HIV/AIDS is a disease of grave global importance with over 33 million people infected world-wide and nearly 2 million deaths each year. The rapid emergence of drug resistance, due to viral mutation, renders anti-retroviral drug candidates ineffective with alarming speed and regularity. Instead of targeting mutation prone viral proteins, an alternative approach is to target host proteins that interact with viral proteins and are critical for the HIV life-cycle. APOBEC3G is a host anti-HIV restriction factor that can exert tremendous negative pressure by hypermutating the viral genome and has the potential to be a promising candidate for anti-retroviral therapeutic research. The work presented in this thesis is focused on investigating the A3G catalytic domain structure and implications of various observed structural features for biological function. High-resolution crystal structures of the A3G catalytic domain were solved using data from macromolecular X-ray crystallographic experiments, revealing a novel intermolecular zinc coordinating motif unique to A3G. Major intermolecular interfaces observed in the crystal structure were investigated for relevance to biochemical activity and biological function. Co-crystallization with a small-molecule A3G inhibitor, discovered using high-throughput screening assays, revealed a cysteine residue near the active site that is critical for inhibition of catalytic activity by catechol moieties. The serendipitous discovery of covalent interactions between this inhibitor and a surface cysteine residue led to further biochemical experiments that revealed the other cysteine, near the active site, to be critical for inhibition. Computational modeling was used to propose a steric-hinderance based mechanism of action that was supported by mutational experiments. Structures of other human APOBEC3 homologs were modeled using in-silico methods examined for similarities and differences with A3G catalytic domain crystal structures. Comparisons based on these homology models suggest putative structural features that may endow substrate specificity and other characteristics to the APOBEC3 family members

Ion-Induced Soot Nucleation Using a New Potential for Curved Aromatics

A potential able to capture the properties and interactions of curved polycyclic aromatic hydrocarbons (cPAHs) was developed and used to investigate the nucleation behaviour and structure of nascent soot particles. The flexoelectric charge polarisation of cPAHs caused by pentagon integration was included through the introduction of off-site virtual atoms, and enhanced dispersion interaction parameters were fitted. The electric polarisation and intermolecular interactions of cPAHs were accurately reproduced compared to ab initio calculations. This potential was used within molecular dynamics simulations to examine the homogeneous and heterogeneous nucleation behaviour of the cPAH corannulene and planar PAH coronene across a range of temperatures relevant to combustion. The enhanced interactions between cPAHs and potassium ions resulted in significant and rapid nucleation of stable clusters compared to all other systems, highlighting their importance in soot nucleation. In addition, the resulting cPAH clusters present morphologies distinct from the stacked planar PAH clusters.This work used the ARCHER UK National Supercomputing Service (http://www.archer.ac.uk). K.B. is grateful to the Cambridge Trust and the Stanley Studentship at King’s College, Cambridge for their financial support. This project is also supported by the National Research Foundation (NRF), Prime Minister’s Office, Singapore under its Campus for Research Excellence and Technological Enterprise (CREATE) programme