This is a revised proposal. The long term goal is to build a comprehensive structural library that will provide information for predictive modeling of three-dimensional structures of nucleic acid sequences of biological significance. The objective is to eventually be able to correlate sequence information and biological function based on known structural modules and motifs. The initial focus is on the triplet repeat (TR) sequences. Spontaneous progressive chain length expansion of these TR sequences has been implicated in more than 10 hereditary neurodegenerative diseases, including fragile X-syndrome, myotonic dystrophy, and Kennedy's disease. The research emphasizes identifying the molecular features and basic properties that differentiate the genetically mutable from non-mutable TR sequences, and the short from long sequences. This information on the structure-based mechanisms for association of TR sequences is aimed at giving possible insights into the link between TR expansion and an hereditary neurodegenderative diseases. Understanding the structure and dynamics of the TR sequences are also anticipated to contribute to the identification of diagnostic and therapeutic targets for TR related genetic diseases. Structure, stability and conformational flexibility of d(CXG)n sequences, as well as the newly discovered d(GAA)n and d(TTC)n sequences, will be studied using NMR spectroscopy under """"""""close to physiological conditions."""""""" (X = A, C, G or T, and n = number of repeats). The NMR information will be correlated with thermodynamics parameters derived from UV measurements.
The specific aims i nclude: 1) Characterization of the sequence alignment and helical conformation of the GAA and TTC TR sequences and the variations of these properties as a function of chain length; 2) Elucidation of the high resolution structures and properties, such as conformation exchange and flexibility, of CAG, CGG, GAC, and new TR sequences that exhibit unusual features; 3) Characterizations of folding topology and folded structures of longer TR sequences; 4) Investigation of the relative stabilities of the homo-duplexes and complementary hetero-duplexes. Studies of structural transitions as a function of chain length.
| Han, X; Gao, X (2001) Sequence specific recognition of ligand-DNA complexes studied by NMR. Curr Med Chem 8:551-81 |
| LeProust, E M; Pearson, C E; Sinden, R R et al. (2000) Unexpected formation of parallel duplex in GAA and TTC trinucleotide repeats of Friedreich's ataxia. J Mol Biol 302:1063-80 |
| Huang, X; Yu, P; LeProust, E et al. (1997) An efficient and economic site-specific deuteration strategy for NMR studies of homologous oligonucleotide repeat sequences. Nucleic Acids Res 25:4758-63 |
