Presentation Title
Structurally Critical Elements for MutY DNA Glycosylase Recognition of Specific Damage Sites
Abstract
Bacteria have evolved a fleet of repair enzymes, in the base excision repair (BER) and GO repair pathways, that combat oxidative DNA damages that have been linked to the initiation and progression of cancer. MutY DNA glycosylases have been shown to locate and remove adenine when incorrectly paired with oxidative 8-oxoguanine (8-OG) damages; failure to remove these damages has been correlated with familial colorectal cancer. In order to investigate the structural features of both the DNA substrate and the MutY glycosylase that allow MutY to find OG:A lesions, we utilize single molecule fluorescence microscopy to observe the enzyme interaction with the substrate in real time. Previous experimentation has suggested the 2-amino group of 8-OG is critical in MutY glycosylase recognition of OG-A lesions. Our goal is to compare MutY interactions with undamaged and OG:A damaged to those with a substrate analog, 8-oxoinosine (8-OI), which lacks this 2-amino group. Diffusive behavior of wild-type MutY on undamaged concatamer showed primarily fast diffusion with few paused or immobile states. Mean squared displacement analysis of the data collected in experimental trials of wild-type MutY on 8-oxoguanine damaged concatamers showed a unimodal distribution indicative of indicative of slow diffusion and significant pausing. Trials are currently being conducted with 8-oxoinosine damaged concatamers. Trials thus far have implied the necessity of the 2-prime amino group for recognition, but 8-oxoinosine data is needed to come to a supported conclusion.
Primary Faculty Mentor Name
Andrea Lee
Status
Undergraduate
Student College
College of Agriculture and Life Sciences
Program/Major
Microbiology and Molecular Genetics
Second Program/Major
Animal Science
Primary Research Category
Biological Sciences
Structurally Critical Elements for MutY DNA Glycosylase Recognition of Specific Damage Sites
Bacteria have evolved a fleet of repair enzymes, in the base excision repair (BER) and GO repair pathways, that combat oxidative DNA damages that have been linked to the initiation and progression of cancer. MutY DNA glycosylases have been shown to locate and remove adenine when incorrectly paired with oxidative 8-oxoguanine (8-OG) damages; failure to remove these damages has been correlated with familial colorectal cancer. In order to investigate the structural features of both the DNA substrate and the MutY glycosylase that allow MutY to find OG:A lesions, we utilize single molecule fluorescence microscopy to observe the enzyme interaction with the substrate in real time. Previous experimentation has suggested the 2-amino group of 8-OG is critical in MutY glycosylase recognition of OG-A lesions. Our goal is to compare MutY interactions with undamaged and OG:A damaged to those with a substrate analog, 8-oxoinosine (8-OI), which lacks this 2-amino group. Diffusive behavior of wild-type MutY on undamaged concatamer showed primarily fast diffusion with few paused or immobile states. Mean squared displacement analysis of the data collected in experimental trials of wild-type MutY on 8-oxoguanine damaged concatamers showed a unimodal distribution indicative of indicative of slow diffusion and significant pausing. Trials are currently being conducted with 8-oxoinosine damaged concatamers. Trials thus far have implied the necessity of the 2-prime amino group for recognition, but 8-oxoinosine data is needed to come to a supported conclusion.