Metal-binding sites in the major groove of a large ribozyme domain.
BACKGROUND
Group I self-splicing introns catalyze sequential transesterification reactions within an RNA transcript to produce the correctly spliced product. Often several hundred nucleotides in size, these ribozymes fold into specific three-dimensional structures that confer activity. The 2.8 A crystal structure of a central component of the Tetrahymena thermophila group I intron, the 160-nucleotide P4-P6 domain, provides the first detailed view of metal binding in an RNA large enough to exhibit side-by-side helical packing. The long-range contacts and bound ligands that stabilize this fold can now be examined in detail.


RESULTS
Heavy-atom derivatives used for the structure determination reveal characteristics of some of the metal-binding sites in the P4-P6 domain. Although long-range RNA-RNA contacts within the molecule primarily involve the minor groove, osmium hexammine binds at three locations in the major groove. All three sites involve G and U nucleotides exclusively; two are formed by G.U wobble base pairs. In the native RNA, two of the sites are occupied by fully-hydrated magnesium ions. Samarium binds specifically to the RNA by displacing a magnesium ion in a region critical to the folding of the entire domain.


CONCLUSIONS
Bound at specific sites in the P4-P6 domain RNA, osmium (III) hexammine produced the high-quality heavy-atom derivative used for structure determination. These sites can be engineered into other RNAs, providing a rational means of obtaining heavy-atom derivatives with hexammine compounds. The features of the observed metal-binding sites expand the known repertoire of ligand-binding motifs in RNA, and suggest that some of the conserved tandem G.U base pairs in ribosomal RNAs are magnesium-binding sites.
Cate
J H
JH
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520, USA.
Doudna
J A
JA
D000818:Animals; D001665:Binding Sites; D003198:Computer Simulation; D003461:Crystallography; D008274:Magnesium / Q000737:chemistry / Q000378:metabolism; D008670:Metals / Q000737:chemistry* / Q000378:metabolism; D019216:Metals, Heavy / Q000737:chemistry / Q000378:metabolism; D008674:Metals, Rare Earth / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009942:Organometallic Compounds / Q000737:chemistry / Q000378:metabolism; D016337:RNA, Catalytic / Q000737:chemistry* / Q000378:metabolism; D016053:RNA, Protozoan / Q000737:chemistry* / Q000378:metabolism; D016808:Tetrahymena thermophila
D008670:Metals; D019216:Metals, Heavy; D008674:Metals, Rare Earth; D009942:Organometallic Compounds; D016337:RNA, Catalytic; D016053:RNA, Protozoan; D008274:Magnesium