X-RAY DIFFRACTION STUDIES OF NUCLEIC ACID STRUCTURES

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The overall objective of this project is to continue investigating the conformational diversities of nucleic acids using x-ray crystallography and nuclear magnetic resonance spectroscopy. Other biophysical techniques, such as laser Raman and circular dichroism spectroscopic methods, will be applied when necessary. Several molecular systems, containing carefully-designed DNA/RNA oligonucleotide sequences associated with important biological implications, will be studied and their structures correlated with biological functions. We propose to study the following DNA/RNA systems for various objectives. (i). Unusual nucleic acid con formations: Several new unusual nucleic acid conformations (e.g., Z-DNA, H-DNA, I-DNA, Hoogsteen helix, telomere, centromere) have been uncovered recently. We will continue to study the molecular basis of those structures and to explore new and unusual conformations. (ii). Nucleic acid structure with defects. Oligonucleotides with different kinds of irregular features such as mismatch, bulge and carcinogen-modified base (e.g., m6G/m4T, benzopyrene-adduct) will be studied. Different modified nucleotide analogs will be incorporated into double helix to assess the effect of them on the stability of various alternative DNA/RNA conformations. (iii). Higher-ordered structures. Nucleic acid molecules can form quite complex tertiary structures like triple helix, hairpin, cruciform, pseudoknot and circular molecules, all of which are important in biological processes (e.g., in gene regulation and recombination). We will continue to focus on the structures of hairpins and circular DNA. (iv). Ligand-nucleic acid interactions. The function of nucleic acids depends on their interactions with other molecules. We will study the interactions between a number of small ligands (in particular metal ions) and nucleic acids by x-ray crystallography. We will explore the crystallization of a few non-sequence-specific DNA binding proteins (e.g., M13 gene V protein and HU) and their DNA complexes. A significant number of those molecules described above are already in various stages of structural analysis. New molecules in the above scope will be synthesized and studied structurally by x-ray diffraction and NMR. The detailed descriptions of various molecular systems are listed in the Preliminary Results section. Our studies will help understand the molecular forces that governs the structure, dynamics and energetics of nucleic acids.