Thermal Analysis of competition between Netropsin and AT Hook Motif binding to DNA

Patrick Lecture Room, Plyler Hall 126

High mobility group A, HMGA, proteins are architectural proteins that bind to the minor groove of AT-rich regions of DNA and can induce conformational changes in DNA. HMGA proteins bind DNA using AT hook motifs, which are composed of RGRP consensus sequences that are typically flanked by basic residues. In adults, overexpression of HMGA proteins is commonly found in a variety of neoplasia. We are investigating how a peptide mimic of the AT hook motif, ATHW, alters Netropsin binding to DNA. Studying the competition between Netropsin and ATHW has implications on the development of compounds that inhibit HMGA proteins. Netropsin is a small polyamide that also binds to AT-rich regions of the minor groove of DNA. Isothermal titration calorimetry, ITC, gives insight on the thermal stability, the water-accessible surface area, and the affect of DNA sequence on ATHW, Netropsin, and the competition between both molecules for binding DNA. ITC results indicate a change in heat capacity, ?Cp, of -0.11 kcal/mol for Netropsin binding to a short DNA containing the AATT sequence. Although further repeats are needed, initial results show a ?Cp of -0.17 kcal/mol for Netropsin competing with ATHW to bind to the an AATT DNA and a ?CP of -0.04 kcal/mol for ATHW binding to AATT DNA. Preliminary work with a DNA sequence containing ATATAT, shows that the ??Cp is not equivalent to the ?Cp of ATHW binding to ATATAT. Thus, the presence of ATHW alters Netropsin binding in a DNA sequence dependent manner.