Self-splicing introns

The discovery of self-splicing introns showed that RNA could catalyse chemical reactions. Yet, unlike proteins, RNA has no functional groups with pKa values and chemical properties similar to those considered to be important in protein-based enzymes. Steitz and Steitz (1993) postulated that two metal ions were essential for catalysis by ribozymes using a mechanism similar to DNA cleavage, in which a free 3 OH is produced. They proposed,... 

As stated previously in the introductory section, T. R. Cech and co-workers reported on the first catalytic RNA or ribozyme, the self-splicing intron of the... 

Another X-ray crystallographic structure, at 3.1-A resolution, of the purple bacterium Azoarcus sp. group I self-splicing intron was published in 2004 by... 

Lehnert, V., Jaeger, L., Michele, F., and Westhof, E. (1996). New loop-loop tertiary interactions in self-splicing introns of subgroup IC and ID A complete 3D model of the Tetrahymena thermophila ribozyme. Chem. Biol. 3, 993—1009. 

In at least one eukaryote, Tetmhymem, the pre-rRNA molecule contains an intron. Removal of the intron during processing of the pre-rRNA does not require the assistance of any protein Instead, in the presence of guanosine, GMP, GDP or GTP, the intron excises itself, a phenomenon known as selfsplicing. This was the first demonstration of ribozymes, that is, catalytic RNA molecules that catalyze specific reactions. The list of ribozymes is growing. For example, self-splicing introns have been discovered in some eukaryotic mRNAs and even peptidyl transferase, a key enzyme activity in protein synthesis, is now known to be a ribozyme (see Topic H2). 

Fig. S. In vitro transcription of a cloned DNA fragment from Anabaena azollae containing a self-splicing intron. (A) Restriction map of a 2.7-kb insert in pBSM13—. Arrows indicate direction of transcription from the T3 and T7 promoters in the vector. tRNA exon (solid) and intron (hatched) sequences are indicated. (B) Plasmid DNA truncated by Ps/I (P), SlpI (S), Dral (D), and Hindi) (H) (in the 3 exon), downstream of the T3 promoter. After transcription with T3 RNA polymerase, products were fractionated on a 3% polyacrylamide-8 M urea gel the autoradiogram is shown. Scale at left indicates position of Haelll restriction fragments of phage 0X174 DNA in nucleotides. Labels indicate positions expected for the unspliced run-off transcript (Pre), ligated exons (LE), and linear intron (LI). (From Xu et at. Copyright 1990 by the AAAS.)...

First, given the wide range of prebiotic nucleic acids, ribose-based polymers may be the most eminently suited for catalysis. Eschenmoser has pointed out, for example, that nucleic acids constructed from hexose nucleotides form inflexible ribbon structures,61 poorly suited for convoluting into the complex shapes that are required for catalysis (e.g., the backbone of the projected tertiary structure of the Tetrahymena self-splicing intron folds back on itself a number of times).62 Conversely, backbones composed of acyclic nucleotides may be too flexible to adopt stable secondary structures (since a great deal of entropy would necessarily be lost on freezing into a given conformer). Ribose, on the other hand, has a limited flexibility because of its pseudorotation cycle, and RNA can adopt a variety of helical conformations. 

Design of RNA molecules with novel catalytic functions called ribozymes (ribonucleotide enzymes) started out from the reprogramming of naturally occurring molecules to accept unnatural substrates [32, 33] A specific RNA cleaving ribozyme, a class I (self-splicing) intron, was modified through variation and selection until it operated efficiently on DNA. The evolutionary path of such a transformation of catalytic activity has been recorded in molecular detail [34]. The basic problem in the evolutionary design of new catalysts is the availability of appropriate analytical tools for the detec-... 

Analysis of Divalent Metal Ion Interactions in the Group I Self-Splicing Intron Active Site... 

Group I (GI) introns are large ribozymes that function as self-splicing intron sequences that catalyze two successive phos-photransesterification reactions using a single conserved active... 

The finding of enzymatic activity in the self-splicing intron and in the RNA component of RNAse P has opened new areas of inquiry and changed the way in which we think about molecular evolution. The discovery that RNA can be a catalyst as well as an information carrier suggests that an RNA world may have existed early in the evolution of life, before the appearance of DNA and protein (Section 2.2.2). 

Figure 28.35. Structure of a Self-Splicing Intron. The structure of a large fragment of the self-splicing intron from Tetrahymena reveals a complex folding pattern of helices and loops. Bases are shown in green, A yellow, C purple, G and orange, U.

The method has also been used to identify residues essential for the splicing activity of self-splicing introns where the spliced molecules are easily separated from unspliced RNA. 

The snRNAs In the spliceosome are thought to have an overall tertiary structure similar to that of group II self-splicing Introns. 

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