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Mycobacterium tuberculosis is an exquisitely adapted intracellular pathogen that encounters hostile, host-derived reactive nitrogen and oxygen intermediates during the course of infection of its human host. These radicals cause DNA damage, which is repaired through various pathways to allow for the continued survival of the organism. Base excision repair (BER) is one such pathway, which depends on DNA glycosylases to identify and excise damaged DNA bases. Formamidopyrimidine DNA glycosylase (Fpg/ MutM/ FAPY) and Endonuclease VIII (Nei) are such enzymes, which both target oxidatively damaged DNA and together, form the Fpg family of DNA glycosylases. Bioinformatic analyses identified two copies each of Fpg and Nei-encoding genes in M. tuberculosis as well as in its non-pathogenic relative, Mycobacterium smegmatis. To understand the role of these multiple glycosylases in the maintenance of genomic integrity and survival of mycobacteria, the genes encoding the four Fpg/Nei glycosylases were individually deleted in M. smegmatis strain mc2155 by homologous recombination. In addition to the four single mutants, double and triple Fpg and Nei glycosylase knockout mutants were generated by sequential gene knockout. When compared to the parental strain, the single and double mutants showed no variation in growth kinetics, no increased sensitivity to hydrogen peroxide and no increase in spontaneous mutation rates. However, a slight increase in frequency of spontaneous C T transition mutations was observed in double knockout mutants compared to the wild type and single mutant strains. These results suggest that these enzymes may be part of an extensive network of enzymes which collectively work to enhance the overall survival of M. smegmatis through the repair of oxidatively damaged DNA.

by Sik Lok Lam. / The "2" in the title is subscript. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1996. / Includes bibliographical references (leaves 184-197). / ABSTRACT --- p.iii / ACKNOWLEDGEMENTS --- p.v / Chapter CHAPTER ONE: --- LITERATURE SURVEY OF SEQUENCE-SPECIFIC LOCAL STRUCTURAL VARIATIONS IN DEOXYRIBONUCLEIC ACID MOLECULES --- p.1 / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.2 --- General Review of DNA --- p.1 / Chapter 1.2.1 --- "Nomenclature, Symbols and Atomic Numbering Scheme of DNA" --- p.2 / Chapter 1.2.2 --- Conformations of DNAs --- p.6 / Chapter 1.2.3 --- Helix-to-Random-Coil Transition --- p.9 / Chapter 1.3 --- Sequence-Specific Local Structural Studies --- p.11 / Chapter 1.4 --- Purpose of This Work --- p.14 / Chapter CHAPTER TWO: --- DETERMINATION OF STRUCTURES OF SOLUTION DNA MOLECULES --- p.17 / Chapter 2.1 --- Introduction --- p.17 / Chapter 2.2 --- Optimization of Conditions --- p.17 / Chapter 2.3 --- Resonance Assignments --- p.19 / Chapter 2.4 --- Extraction of Structural Constraints --- p.22 / Chapter 2.4.1 --- Interproton Distances --- p.23 / Chapter 2.4.2 --- Endocyclic Sugar Torsion Angles --- p.25 / Chapter 2.4.3 --- Phosphate Backbone Torsion Angles --- p.29 / Chapter 2.4.4 --- Hydrogen Bonds --- p.31 / Chapter 2.5 --- Structural Refinement --- p.31 / Chapter CHAPTER THREE: --- SIGNIFICANCE OF DIFFERENT TYPES OF STRUCTURAL CONSTRAINTS IN STRUCTURAL REFINEMENT PROCESS --- p.35 / Chapter 3.1 --- Introduction --- p.35 / Chapter 3.2 --- Experimental --- p.36 / Chapter 3.2.1 --- DNA Model Building --- p.36 / Chapter 3.2.2 --- Generation of Structural Constraints --- p.37 / Chapter 3.2.3 --- Structural Refinement --- p.40 / Chapter 3.3 --- Results and Discussion --- p.41 / Chapter 3.3.1 --- Endocyclic Sugar Torsion Angle Constraints --- p.45 / Chapter 3.3.2 --- Phosphate Backbone Torsion Angle Constraints --- p.49 / Chapter 3.3.3 --- Hydrogen Bond Constraints --- p.50 / Chapter 3.4 --- Summary --- p.50 / Chapter CHAPTER FOUR: --- EFFECTS OF DIFFERENT VARIABLES IN THE RESTRAINED MOLECULAR DYNAMICS PROCESS --- p.52 / Chapter 4.1 --- Introduction --- p.52 / Chapter 4.2 --- Experimental --- p.53 / Chapter 4.3 --- Results and Discussion --- p.55 / Chapter 4.3.1 --- Variables in the Temperature Profile --- p.58 / Chapter 4.3.2 --- Variables in the Force Constant Profile --- p.62 / Chapter 4.4 --- Summary --- p.65 / Chapter CHAPTER FIVE: --- THE J-COUPLING RESTRAINED MOLECULAR MECHANICS PROTOCOL - AN EFFICIENT AND RELIABLE ALTERNATIVE IN DERIVING ENDOCYCLIC SUGAR TORSION ANGLE CONSTRAINTS --- p.66 / Chapter 5.1 --- Introduction --- p.66 / Chapter 5.2 --- Methodology --- p.71 / Chapter 5.2.1 --- "Establishment of the Correlation of 3J1'2, withvi" --- p.71 / Chapter 5.2.2 --- Sample Preparation --- p.73 / Chapter 5.2.3 --- NMR Analysis --- p.73 / Chapter 5.2.4 --- Theoretical Testing of the Protocol --- p.74 / Chapter 5.2.5 --- Experimental Testing of the Protocol --- p.75 / Chapter 5.3 --- Results and Discussion --- p.76 / Chapter 5.3.1 --- Selection of the Appropriate JrMM-derived Torsion Angles --- p.85 / Chapter 5.3.2 --- Theoretical Testing of the Protocol --- p.88 / Chapter 5.3.3 --- Experimental Testing of the Protocol --- p.93 / Chapter 5.4 --- Summary --- p.98 / Chapter CHAPTER SIX: --- HETERONUCLEAR SINGLE QUANTUM COHERENCE DERIVED BACKBONE TORSION ANGLE CONSTRAINTS --- p.99 / Chapter 6.1 --- Introduction --- p.99 / Chapter 6.2 --- Experimental --- p.102 / Chapter 6.3 --- Results and Discussion --- p.103 / Chapter 6.3.1 --- Determination of the Backbone Torsion Angles β and E --- p.103 / Chapter 6.3.2 --- Error Estimation on 3JC4'p- and 3JH3'p-derived E --- p.109 / Chapter 6.4 --- Summary --- p.110 / Chapter CHAPTER SEVEN: --- SOLUTION STRUCTURES OF d(CGXX，CG)2 AND d(CAXX´ةTG)2 --- p.111 / Chapter 7.1 --- Introduction --- p.111 / Chapter 7.2 --- Experimental --- p.111 / Chapter 7.2.1 --- Sample Preparation --- p.112 / Chapter 7.2.2 --- Resonance Assignment --- p.112 / Chapter 7.2.3 --- Melting Profile Study --- p.112 / Chapter 7.2.4 --- Extraction of Structural Constraints --- p.113 / Chapter 7.2.5 --- Structural Refinement --- p.115 / Chapter 7.2.6 --- Structural Parameter Analysis --- p.116 / Chapter 7.3 --- Results and Discussion --- p.116 / Chapter 7.3.1 --- Melting Profile Study --- p.117 / Chapter 7.3.2 --- Structural Constraints --- p.120 / Chapter 7.3.3 --- Structural Refinement --- p.129 / Chapter 7.3.4 --- Structural Features --- p.135 / Chapter CHAPTER EIGHT: --- SEQUENCE-SPECIFIC LOCAL STRUCTURAL STUDY --- p.156 / Chapter 8.1 --- Introduction --- p.156 / Chapter 8.2 --- Predictions from the Calladine's Rules --- p.156 / Chapter 8.3 --- Predictions from Olson's Base-Pair Morphology Dependent Clash Function --- p.160 / Chapter 8.4 --- Re-formulation of Calladine's Idea and its Relationship to Sequence-Specific Local Structural Function ΣLS --- p.163 / Chapter 8.4.1 --- Sequence-Specific Base-Pair Geometry Analysis --- p.164 / Chapter 8.4.2 --- Sequence-Specific Base-Pair Step Geometry Analysis --- p.166 / Chapter 8.4.3 --- Sequence-Specific Local Structural Function ΣLS --- p.167 / Chapter 8.5 --- Summary --- p.173 / Chapter CHAPTER NINE: --- CONCLUSIONS AND FURTHER WORK --- p.174 / APPENDIX I The Base Proton Regions of the lH NMR Spectra of the Hexamers --- p.177 / APPENDIX II 2D NOESY Spectra (Tm = 200 ms) of the Hexamers --- p.178 / "APPENDIX III The H1'-H27H2"" Regions of the DQF-COSY Spectra of the Hexamers" --- p.180 / APPENDIX IV The C4'-H4' Regions of the HSQC Spectra of the Hexamers --- p.182 / REFERENCES --- p.184

The extent of genetic structuring of a population results from a balance of forces producing local genetic differentiation, and counter-forces producing genetic homogeneity. An understanding of these forces is essential when investigating evolutionary processes in a species. It has been predicted that, when populations experience severe reductions in size, they lose genetic variability. Small population size may occur as a consequence of founder events, such as with the introduction and spread of a pest species. The predictions as described were examined by studying the molecular evolution and population genetics of European rabbit (Oryctolagus cuniculus (L)) populations in Australia, using variation in mitochondrial DNA control region sequences. The results suggest that there is a higher rate of gene flow within populations in semi arid/arid areas, which is attributable to the extreme environmental heterogeneity found in these areas and may be best explained by a process whereby local populations becoming extinct and are recolonised by individuals drawn from other populations. / Master of Science (Hons)

Graduation date: 2003

Thesis (M.S. in biochemistry)--Washington State University, December 2009. / Title from PDF title page (viewed on Jan. 20, 2010). "School of Molecular Biosciences." Includes bibliographical references.

Thesis (Ph. D.)--University of Texas at Austin, 1999. / Vita. Includes bibliographical references (leaves 202-222). Available also in a digital version from Dissertation Abstracts.

Thesis (Honors)--College of William and Mary, 2009. / Includes bibliographical references (leaves 51-54). Also available via the World Wide Web.

Thesis (Ph. D.)--University of Texas at Austin, 2001. / Vita. Includes bibliographical references (leaves 144-154). Available also in a digital version from Dissertation Abstracts.