It is evident from many different experiments, that the human nucleotide excision repair (NER) apparatus first distinguishes structural DNA perturbations caused by bulky lesions derived from the reactions of metabolically activated environmental carcinogens that enter the human body. This step entails the formation of XPC/HR23B complexes that partially open the duplex near the lesion site. The initial structural distortions caused by the lesions and the subsequent strand separation suggest that base-base stacking and hydrogen bonding interactions are first weakened and then broken during these initial phenomena. In this project, we will examine the effects of structurally different lesions of different adduct conformations on these initial structural distortions, and then use variations in base sequence context as a tool to modulate these base stacking interactions and thus gain insight into the specific structural factors that affect human NER activity.
In Specific Aim 1, the Structural basis of recognition and processing of DNA damage by the human NER apparatus will be investigated.
This aim will be focused on DNA lesions of different chemical structure, physical size, and conformational properties positioned at the same site in otherwise completely identical sequence contexts.
In Specific Aim 2, the local DNA distortions caused by the lesions will be modulated by varying the base sequence context in which the lesions are embedded, and determine effects of different flanking bases on the structural characteristics and changes in NER activity.
In Specific aim 3, the effects of adduct structure and base sequence on binding and patterns of helix opening by the human NER lesion-recognizing XPC/HR23B heterodimer duplex will be investigated. The results of this project will have ultimate translational identification by providing new information about DNA lesiosn that are resistant to DNA repair, thus providing important information about biomarkers of exposure of the human population to environmental carcinogens.
Exposure of the human population to cancer-causing substances in the environment, constitutes an important hazard to human health. Among these chemicals are polycyclic aromatic hydrocarbons in urban polluted environments and in cigarette smoke, and aromatic amine mutagens present in cooked foods. These chemicals, once they enter the human body, are activated to highly reactive intermediates that chemically bind to DNA, thus generating bulky adducts. The latter, if not removed by cellular defense mechanism, can cause mutations and cancer. One of the critical lines of defense of the human body to these environmental carcinogens is DNA repair, specifically by the nucleotide excision repair (NER) mechanism that deals with bulky DNA damage. Unfortunately, the molecular basis of this critical defense mechanism, especially its efficiency in removing important, structurally different DNA lesions, is still obscure. In this project, a systematic approach towards solving this problem is proposed that will identify the kind of carcinogen-DNA lesions that are poorly repaired by NER mechanisms. This information will be useful in biomarker studies of environmental human exposure, and will be especially useful for identifying individuals who, because of genetic polymorphisms, are of particular risk of developing cancer. Such individuals can then be selected for closer monitoring to prevent the development of this disease into its later life-threatening stages.
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