Sequence motifs and free energies of selected natural and non-natural nucleosome positioning DNA sequences

Our laboratories recently completed SELEX experiments to isolate DNA sequen ces that most-strongly favor or disfavor nucleosome formation and positioni ng, from the entire mouse genome or from even more diverse pools of chemica lly synthetic random sequence DNA. Here we directly compare these selected natural and non-natural sequences. We find that the strongest natural posit ioning sequences have affinities for histone binding and nucleosome formati on that are sixfold or more lower than those possessed by many of the selec ted non-natural sequences. We conclude that even the highest-affinity seque nce regions of eukaryotic genomes are not evolved for the highest affinity or nucleosome positioning power. Fourier transform calculations on the sele cted natural sequences reveal a special significance for nucleosome positio ning of a motif consisting of similar to 10 bp periodic placement of TA din ucleotide steps. Contributions to histone binding and nucleosome formation from periodic TA steps are more significant than those from other periodic steps such as AA (=TT), CC (=GG) and more important than those from the oth er YR steps (CA (=TG) and CG), which are reported to have greater conformat ional flexibility in protein-DNA complexes even than TA. We report the deve lopment of improved procedures for measuring the free energies of even stro nger positioning sequences that may be isolated in the future, and show tha t when the favorable free energy of histone-DNA interactions becomes suffic iently large, measurements based on the widely used exchange method become unreliable. (C) 1999 Academic Press.