Experimental and computational studies of DNA structure using the hydroxyl radical as a chemical probe

Abstract

We have constructed a database of hydroxyl radical (•OH) cleavage patterns of DNA in order to investigate the relationship between the sequence of a DNA molecule and its three-dimensional structure. The hydroxyl radical cuts DNA at every nucleotide, with the amount of cutting proportional to the solvent accessible surface area (SASA) of the deoxyribose hydrogen atoms. Cleavage fragments are quantified by a fluorescence sequencer, followed by normalization and deposition into the database. Our database currently contains 151 DNA sequences with lengths ranging from 35 to 41 nucleotides. These data have enabled us to develop some general rules regarding the sequence-dependence of DNA structure as well as to predict the cleavage pattern of any given DNA sequence with remarkable precision. Using this prediction algorithm, it is possible to construct structural maps of entire genomes. As there are many examples of DNA binding proteins with highly degenerate binding sites, the use of structural information to locate these sites may be helpful. There also exists other signals, including the signal for nucleosome positioning, which have no apparent consensus, making it likely that the structure of DNA is of critical importance. We have developed algorithms to identify regions of conserved structure using •OH cleavage intensity as a proxy. Within a set of DNase I hypersensitive sites (DHS) obtained from the ENCODE Consortium, we were able to identify a stretch of 12 nucleotides for which the structural conservation is much greater than the sequence conservation. These sites have been dubbed Conserved •OH Radical Cleavage Signatures, or CORCS. Upon further analysis, these CORCS were found to be 17-fold enriched for DHS as compared to shuffled elements. Through the continued analysis of hydroxyl radical cleavage data and development of algorithms to employ the data in biologically meaningful ways, we hope to further our understanding of the relationship between DNA sequence and structure, and how the local structural heterogeneity of genomic DNA contributes to biological function.