The conformation of chicken erythrocyte chromatin multimers will be determined from the molecular weight dependence of the friction factor ratio f/fo, obtained by quasielastic light scattering (QLS) and sedimentation velocity experiments. Once the values of D and S, hence Mr, are known for a homologous series of chromatin multimers (i.e., with and without the lysine-rich histones H1 and H5 for chicken erythrocytes), then the Kirkwood theory or the Bloomfield-Dalton-Van Holde model for hydrodynamic interaction between friction beads can be used to simulate the data, hence inference of the molecular conformation in solution. The initial series of experiments will be directed to the parameters describing the basic unit of all supramolecular structures, i.e., the dimer. The equivalent hydrodynamic radius of the "core particle" in the Van Holde model for chromatin substructure, or the "nu body", will be determined from the purified monomer pool. Once this value is known precisely and accurately, purified dimers will be studied to determine the hydrodynamic distance between two core particles. These studies will be carried out as a function of ionic strength, temperature, effect of lysine-rich histones, and response to the presence of intercalating molecules such as psoralen in an attempt to characterize completely the "spacer region" postulated by Van Holde, et al. This concept has chemical support but, until recently, as described herein, has found no support by physical measurements . After complete characterization of the basic unit, the supramolecular structure for chromatin multimers will be similarly characterized. The popular conformation is a helical structure with 4-10 "beads" per turn, although other conformations may be equally likely, i.e., flexible coil with attractive interactions between beads. The ultimate goal of the present proposal is to determine uniquely the conformation of chromatin multimers, with and without lysine-rich histones, in solution. Completion of this project would represent the first detailed study of the conformation of chromatin in solution, which, hopefully, will provide invaluable hydrodynamic information which may relate to chromatin condensation.
NIH ExPorter
NIH ExPorter