Computational studies of damaged DNA : an investigation of DNA O-linked adducts formed due to exposure to phenolic carcinogens

This thesis systematically develops a computational model to identify the conformational and base-pairing preferences of PhOdG, 4-Cl-PhOdG, DCP-OdG, TCP-OdG, and PCP-OdG by gradually increasing the size of the system also structural properties of unsubstituted O-linked. All adducts at nucleoside level adopted syn conformation. Moreover, effect of protonation at N3 and N7 site on the structural properties and deglycosilation barrier of adducted guanosine was probed. It was highly desirable to include O-linked phenolic as well as C8-dG adducts into a DNA strand in order to understand the detrimental effect of them and the conformational distortion of double helix duplex the desired modified base into NarI DNA duplex through the employment of molecular dynamic simulation (MD) was assessed. The anti-conformation against cytosine is preferred with this model for all adducts and syn conformer for all unsubstituted O-linked and ortho and para Clinked structures against guanine mismatch is the lowest energy structure.University of Lethbridg

UNDERSTANDING ALLOSTERIC INHIBITION OF DIHYDRODIPICOLINATE SYNTHASE FROM CAMPYLOBACTER JEJUNI

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Influence of the Linkage Type and Functional Groups in the Carcinogenic Moiety on the Conformational Preferences of Damaged DNA: Structural and Energetic Characterization of Carbon- and Oxygen-Linked C⁸‑Phenolic-Guanine Adducts

Computational (DFT, MD, and free energy) methods are used to systematically compare the structural and energetic properties of C⁸-bonded 2′-deoxyguanosine (dG) adducts derived from phenolic toxins, namely, the oxygen-linked (unsubstituted) adduct (^PhOdG) and carbon-linked adducts (^{<i>ortho</i>‑PhOH}dG or ^{<i>para</i>‑PhOH}dG) that contain a hydroxyl group in the bulky moiety. Despite restricted rotation at the C⁸–X bond due to the presence of the oxygen linker, the ^PhOdG adduct likely possesses the greatest glycosidic (<i>anti</i>/<i>syn</i>) conformational flexibility at the 5′-terminus of DNA. However, the <i>anti</i>/<i>syn</i> energy difference is the smallest for the ^{<i>para</i>‑PhOH}dG nucleotide at other helical positions, which correlates with the greatest conformational heterogeneity for the corresponding (<i>Nar</i>I) adducted DNA. Most importantly, the total number of accessible conformations of adducted DNA depend on the phenolic adduct considered. Specifically, although the only conformations accessible to ^PhOdG adducted DNA correspond to the <i>anti</i> adduct glycosidic orientation, the C-linked adducts can also adopt the <i>syn</i> orientation in the double helix. Moreover, the number of accessible conformations for DNA containing the C-linked adducts depends on the nature of discrete interactions involving the hydroxyl group in the C⁸-moiety. In fact, such interactions lead to a novel (intercalated) conformational theme in the case of the ^{<i>para</i>‑PhOH}dG adduct. Together, these results indicate that the type of C⁸-linkage, and the presence and location of additional functional groups in the bulky moiety affect the conformational outcomes, which adds to the list of previously established effects including the size of the carcinogenic moiety, adduct ionization state, and sequence context on the conformational preferences of damaged DNA. Most importantly, our study provides valuable structural information that explains the experimentally observed mutagenic potential of DNA phenolic adducts and predicts the relative repair propensity of the three phenolic lesions

Structural Influence of C8-Phenoxy-Guanine in the <i>Nar</i>I Recognition DNA Sequence

Phenoxyl radicals can covalently attach to the C8 site of 2′-deoxyguanosine (dG) to generate oxygen-linked biaryl ether C8-dG adducts. To assess the structural impact of an O-linked C8-dG adduct in duplex DNA, C8-phenoxy-G (^PhOG) and C8-4-fluorophenoxy-G (^4FPhOG) were incorporated into the G³ position of the 12-mer <i>Nar</i>I recognition sequence (5′-CT­C­G­G­C­X­C­C­A­TC, where X = G, ^PhOG, or ^4FPhOG) using solid-phase DNA synthesis with O-linked C8-dG phosphoramidites. The modified strands were hybridized to six different complementary strands that include regular base pairing to C [<i>Nar</i>I′(C)], mismatches with G, A, T [<i>Nar</i>I′(N)], and an abasic site [<i>Nar</i>I′(THF)], and a 10-mer sequence to model a −2 deletion duplex [<i>Nar</i>I′(−2)]. All duplex structures were characterized using UV–vis thermal melting temperature analysis, and in each instance, the O-linked C8-phenoxy-G adducts were found to destabilize the duplex relative to the unmodified controls. The most stable duplex structures match the O-linked C8-dG adduct against C and a G mismatch, which are comparable in terms of stability. These duplexes were further characterized using circular dichroism, dynamic ¹⁹F nuclear magnetic resonance experiments, and molecular dynamics simulations. On the basis of these findings, ^PhOdG adopts the B conformation opposite C, with the phenoxy moiety residing in the solvent-exposed major groove. However, opposite the G mismatch, ^PhOdG adopts a “W-type” wedge conformation with the phenoxy group residing in the minor groove. These studies predict that the O-linked C8-dG lesion ^PhOG will have a weak mutagenic effect, as determined for the corresponding single-ringed nitrogen-linked C8-dG adduct derived from aniline

Influence of Chlorine Substitution on the Hydrolytic Stability of Biaryl Ether Nucleoside Adducts Produced by Phenolic Toxins

A kinetic study is reported for the acid-catalyzed hydrolysis of oxygen (<i>O</i>)-linked biaryl ether 8-2′-deoxyguanosine (dG) adducts produced by phenolic toxins following metabolism into phenoxyl radical intermediates. Strikingly, the reaction rate of hydrolysis at pH 1 decreases as electron-withdrawing chlorine (Cl) substituents are added to the phenoxyl ring. The Hammett plot for hydrolysis at pH 1 shows a linear negative slope with ρ_X = −0.65, implying that increased Cl-substitution diminishes the rate of hydrolysis by lowering N⁷ basicity. Spectrophotometric titration provided an N⁷H⁺ p<i>K</i>_a value of 1.1 for the unsubstituted adduct 8-phenoxy-dG (Ph-O-dG). Model pyridine compounds suggest N⁷H⁺ p<i>K</i>_a values of 0.92 and 0.37 for 4-Cl-Ph-O-dG and 2,6-dichloro-Ph-O-dG (DCP-O-dG), respectively. Density functional theory (DFT) calculations also highlight the ability of the 8-phenoxy substituent to lower N⁷ basicity and predict a preference for N³-protonation for highly chlorinated <i>O</i>-linked 8-dG adducts in water. The calculations also provide a rationale for the hydrolytic reactivity of <i>O</i>-linked 8-dG adducts in the gas-phase, as determined using electrospray mass spectrometry (ESI-MS). The inclusion of our data now establishes that the order of hydrolytic reactivity at neutral pH for bulky 8-dG adducts is <i>N</i>-linked > <i>C</i>-linked > <i>O</i>-linked, which correlates with their relative ease of N⁷-protonation