The RNA World Hypothesis -cofactors -ancient enzymes -ribozymes Ribozymes Catalytic RNA Nobel prize - Chemistry 1989
T.R. Cech (Colorado, HHMI) S. Altman (Yale) RNase P Tetrahymena group I Ribozymes Catalytic RNA Catalysis Auto-catalysis -rate enhancement
-turnover -rate enhancement on self reaction Small: hammerhead hepatitis d hairpin VS Medium: group I group II RNase P Large: ribosome spliceosome? Catalysis Acceleration Catalysis Turnover RNA Catalysis How fast? pre-rRNA processing -In eukaryotes, the gene is not continuous..."functional" sequences are interrupted "non-functional" sequences. -First discovered for adenovirus pre-mRNAs (PA Sharp 1977) -Intron/Exon structure (W. Gilbert) of eukaryotic genome Tetrahymena rRNA contains one intron (IVS).How is it removed? Exon Exon Intron Tetrahymena intron removal
Two sequential transesterifications 5 splice site 3 5 3 5 5 5 3 Stereochemical Course of Splicing
-transesterification reactions of splicing are nucleophilic substitution reactions Recall: inversion mirror images R S Inversion Retention Prochiral Centre Made Chiral by S Substitution
5 P P P 3 Rp Sp Stereochemistry of Group I Splicing
-Rp at 5 splice site gives Sp in product -Sp at 3 splice site gives Rp in product Stereochemistry of Group I Splicing
Rp Rp Sp Sp Tetrahymena intron removal
3 5 Nu LG 3 5 5 5 3 One Active Site in Group I Intron
first step second step G Binding Site - Group I Intron
C311 G G G264 G Specificity Swap at G Binding Site - Group I Intron
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X Cell (1998) 92: Altered Hammerhead Conformation Altered Hammerhead Conformation
Molecular Cell, Volume 23,4, ,2006 Altered Conformation: Active Site
Cell, 126, 2, , 2006 Hepatitis d Ribozyme Hepatitis d X-ray Nature (1998) 395:567-74 Hepatitis d X-ray Nature (1998) 395:567-74 General Base Catalysis in a Ribozyme
Science (1999) 286: Imidazole Rescue of Ribozyme Activity
Science (1999) 286: Imidazole Rescue of Ribozyme Activity
Science (1999) 286: