Faculty of Pharmaceutical Sciences, Chiba Institute of Science
○Toshiyuki Chatake Osamu Matsumoto Tatsuhiko Kikkou
The combination of the development of chemical synthesis method for oligonucleotides and the improvement of the X-ray diffraction method has achieved over 500 X-ray structure analyses of nucleic acids in the present two decades. Nevertheless, obtaining single suitable crystals of oligonucleotides is still an empirical technique. During the crystallization, we need to find out the most suitable combination of metal salts and an alcohol as precipitant, and adjust the concentrations of selected chemical reagents to grow better and larger crystals. These steps are trial-and-error procedures, so that they would sometimes require a large effort or end in failure. As the result, a considerable number of three dimensional structures of important nucleic acids haven't been revealed yet.
In this presentation we propose a simple temperature-control method for obtaining single DNA crystals. Two complementary DNA oligomers usually form a double-helical structure by the Watson-Crick base pairing, called as double stranded DNA (dsDNA). Once the temperature rises, these dsDNA are dissociate to be two single stranded DNAs (ssDNA). Since the phosphate groups in ssDNA are exposed into the solvent region, ssDNA is more water soluble than dsDNA. Such change of solubility in the transition from dsDNA to ssDNA is utilized in the present method. In the present investigation, the melting point (Tm) of DNA oligomers in the crystallization solution were estimated to be in the range of 333 - 318 K, and the single crystals of these DNA molecules could be obtained by systematically decreasing from 333 to 293 K.