NSF Award
0616005
The research focuses on the thermodynamics of nucleic acid intramolecular structures - especially, triplet helices because of their implication in the control of cellular processes by endogenous or exogenous mechanisms. The long term objectives are to understand the molecular forces controlling the overall stability of complex intramolecular DNA structures; to quantify the energetics and hydration contributions governing the association of triplexes, and other unusual structures, with their complementary strands, including the role of sequence and cations; and to determine the thermodynamics for their favorable interaction with polycations for cellular delivery purposes. UV and CD spectroscopies are used to verify that each nucleic acid complex contains the appropriate structural features. Further, a combination of T-dependent UV spectroscopy, differential scanning and pressure perturbation calorimetries will be employed to obtain complete thermodynamic profiles for their unfolding reactions as a function of salt, pH and osmolyte concentration. High sensitivity titration calorimetry and density techniques will be used to measure the heat and volume change of association reactions. The complete thermodynamic characterization of these DNA complexes will provide a fundamental understanding of the physical factors that determine their stability as a function of its sequence and solution conditions. These factors are basic to the rational design of gene-targeting reagents, and for their proper cellular delivery, which can be used in therapeutic, diagnostic and biotechnological applications, and for predicting the energetics of sequence-specific local conformational rearrangements in intracellular processes.<br/><br/>The broader impacts of this research are in carcinogenesis and gene therapy because of the fundamental importance of developing highly specific and stable agents for targeting oncogenes or their transcribed RNA products via triplex or duplex formation. Another impact is the role of water in the overall properties of biological macromolecules and in their interacting behavior towards one another. Specifically, the correlation of energetics with hydration will improve the picture of how hydration controls the stability, conformation and melting behavior of these novel nucleic acid structures. Furthermore, the resulting hydration data can be used in molecular modeling studies and in theoretical calculations, providing an insight into global water. The educational significance is in the mentoring of undergraduate, graduate students and postdoctoral fellows by training them with a wide variety of biophysical techniques and with the fundamental understanding of biophysics. In addition, the research findings are routinely incorporated into lectures of undergraduate, graduate courses and seminars.
