The P4-P6 domain directs higher order folding of the Tetrahymena ribozyme core.
The active site of group I self-splicing introns occurs at the interface of two proposed structural domains. In the Tetrahymena intron, half of the catalytic core resides within the independently-folding P4-P6 domain while the other half belongs to a putative domain that includes helices P3, P7, P8, and P9 (P3-P9). To determine whether the P3-P9 region of the intron can also fold independently, we used Fe(II)-EDTA and dimethyl sulfate to probe the solvent accessibility of separate fragments of the Tetrahymena intron. These RNAs self-assemble into an active complex in trans, enabling analysis of their structural features both alone and within the complex. Our results show that while the P3-P9 region of the intron retains its secondary structure, most of the tertiary interactions within this region do not form stably in the absence of the P4-P6 domain. This indicates that the P4-P6 domain induces folding in the P3-P9 region, organizing the catalytic cleft between them. Thus the P4-P6 domain provides a scaffold for the folding of the Tetrahymena intron core.
Doherty
E A
EA
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520, USA.
Doudna
J A
JA
D000818:Animals; D001483:Base Sequence; D004492:Edetic Acid; D005296:Ferrous Compounds; D017125:Genes, Protozoan; D007438:Introns; D008961:Models, Structural; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010957:Plasmids; D016133:Polymerase Chain Reaction; D016337:RNA, Catalytic / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D016053:RNA, Protozoan / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012997:Solvents; D013463:Sulfuric Acid Esters; D013698:Templates, Genetic; D013768:Tetrahymena / Q000235:genetics*
D005296:Ferrous Compounds; D016337:RNA, Catalytic; D016053:RNA, Protozoan; D012997:Solvents; D013463:Sulfuric Acid Esters; C055526:Fe(II)-EDTA; D004492:Edetic Acid; C007482:dimethyl sulfate