P74 Personalizing a large trial: Every sites' contribution counts

Abstract

This chapter discusses the current status, of using nuclear magnetic resonance (NMR), to study the structure and dynamics of the holliday junction (HJ). Complex deoxyribonucleic acid (DNA) structures (e.g., triplexes, quadruplexes, junctions) pose difficult problems for study, by NMR, relative to the typical DNA duplexes, because they have nonstandard or distorted local conformations and higher molecular weights that give rise to large resonance linewidths and severe 1H spectral overlap. With more atoms in the system, both assignment and structure calculation become more challenging. The HJ, a four-arm DNA crossover structure, is a transient intermediate formed in the course of genetic recombination as well as during other cellular processes, such as replication and telomere resolution. A significant body of evidence has accumulated, indicating that the structure at the junction has a central role, in determining the outcome of these cellular events. For NMR studies, the titration of the four component 16-mer strands to create an equimolar mixture is critical. Gel electrophoresis has shown that titrations based on the standard ultraviolet (UV) estimates of strand concentrations result in significant amounts of residual single-strand, half-complementary duplex, and three-arm structures.