Rapid crystallization of chemically synthesized hammerhead RNAs using a double screening procedure.
To find conditions for obtaining diffraction-quality crystals of a hammerhead RNA rapidly and reproducibly, we employed a "double screening" procedure in which we screened six different RNA synthetic constructs against 48 crystallization conditions using a newly devised sparse matrix. We obtained crystals immediately and diffraction-quality crystals of the sixth RNA construct within six months of initiating the screening of additional RNA sequences. The best crystals diffract to 2.9 A resolution when flash-cooled at synchrotron X-ray sources. Solid-support chemical synthesis combined with sparse matrix screening should allow rapid production of diffraction-quality crystals of a variety of small RNAs, reducing the time commitment for initiating such crystallography projects from several years to several months. The synthetic approach also makes introduction of modified bases to prevent self-cleavage and to generate isomorphous heavy-atom derivative crystals a rapid and straightforward process.
Scott
W G
WG
MRC Laboratory of Molecular Biology, Cambridge, England.
Finch
J T
JT
Grenfell
R
R
Fogg
J
J
Smith
T
T
Gait
M J
MJ
Klug
A
A
D001483:Base Sequence; D003460:Crystallization; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D016337:RNA, Catalytic / Q000138:chemical synthesis* / Q000737:chemistry*
D016337:RNA, Catalytic