Conformational Stability & Dynamics of G-Quadruplexed DNA & Ligand Interactions

Brooklyn College, New York NY

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Abstract

Guanine-rich DNA tandem repeat telomeric sequences located at the end of the chromosomes can assume[unreadable] highly stable G-quadruplex structures through Hoogsteen base pairing of the guanine residues. These[unreadable] secondary DNA structures can inhibit the activity of telomerase, an enzyme that is important for[unreadable] tumorigenesis. There is intense interest in developing so-called G-quadruplex interactive agents (QIAs),[unreadable] which are able to stabilize the G-quadruplex structure as potential chemotherapeutics. However,[unreadable] advancement of this promising area of research has been hindered by a general lack of knowledge of the[unreadable] fundamental rules that govern G-quadruplex formation, conformation and dynamics, properties which can[unreadable] influence productive and selective QIA ligand interactions. Hence there a critical need for understanding the[unreadable] basic chemical and physical characteristics of the G-quadruplex. The aims of this research application are:[unreadable] (1) to examine the conformational stability at specific sites within the G-quadruplex and to assess the effects[unreadable] of QIA binding on these parameters; and 2) to examine the underlying dynamics of intramolecular G-quadruplex[unreadable] folding and unfolding. To achieve these aims we have prepared a series of fluorescently labeled[unreadable] human telomeric sequences, with replacement of a single guanine residue by a fluorescent nucleoside[unreadable] analog at varying postions within the G-quadruplex. The conformation and dynamics of the G-quadruplex will[unreadable] be studied using state-of-the-art fluorescence methodologies. Isothermal titration calorimetry (ITC) will be[unreadable] used to examine thermodynamic quantities involved in QIA ligand binding; and electrophoresis and UV[unreadable] analyses to confirm quadruplex formation. The information provided by the combination of these[unreadable] characterization methods will allow us to map the dynamic structure of the quadruplex and determine the[unreadable] impact of QIA binding. The broad, long-term objective of the proposed research is to understand the[unreadable] fundamental "rules" that govern the conformation and dynamics of G-quadruplexed DNA, and to begin to[unreadable] understand the effects of specific ligand binding on them for ultimate applications in the rational design of[unreadable] QIAs.