RAPID CRYSTALLIZATION OF CHEMICALLY SYNTHESIZED HAMMERHEAD RNAS USINGA DOUBLE SCREENING-PROCEDURE

To find conditions for obtaining diffraction-quality crystals of a ham merhead RNA rapidly and reproducibly, we employed a ''double screening '' procedure in which we screened six different RNA synthetic construc ts against 48 crystallization conditions using a newly devised sparse matrix. We obtained crystals immediately and diffraction-quality cryst als of the sixth RNA construct within six months of initiating the scr eening of additional RNA sequences. The best crystals diffract to 2.9 Angstrom resolution when flash-cooled at synchrotron X-ray sources. So lid-support chemical synthesis combined with sparse matrix screening s hould allow rapid production of diffraction-quality crystals of a vari ety of small RNAs, reducing the time commitment for initiating such cr ystallography projects from several years to several months. The synth etic approach also makes introduction of modified bases to prevent sel f-cleavage and to generate isomorphous heavy-atom derivative crystals a rapid and straightforward process.