Self-splicing

Several features of these RNA enzymes, or ribozymes, lead to the realization that their biological efficiency does not challenge that achieved by proteins. First, RNA enzymes often do not fulfill the criterion of catalysis in vivo because they act only once in intramolecular events such as self-splicing. Second, the catalytic rates achieved by RNA enzymes in vivo and in vitro are... 

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... 

Figure 6.7 A bacterial self-splicing group I intron with both exons (PDB 1U6B). Visualized using CambridgeSoft Chem3D Ultra 10.0 with notations in ChemDraw Ultra 10.0.(Printed with permission of CambridgeSoft Corporation.) (See color plate)...

Enzymatically active ribonucleic acid segments (some of which are known as ribozymes) with the capacity to catalyze RNA self-splicing or peptide bond formation. The overall catalytic rate enhancement is around 10 ... 

RNA molecules that undergo self-splicing, in which an internal portion of the RNA molecule is removed while the parts on either side of this intron are reconnected (see Chapter 11). 

Other RNA molecules that do not undergo self-splicing can act on other molecules as substrates are true catalysts. 

Based on the observation of self splicing by the 23S RNA of Tetrahymena, it is assumed that the cleavage and rejoining of the phosphodiester bond is catalyzed by the RNA components of the spliceosome. The proteins of the spliceosome are believed to be important for the recognition of the 5 and 3 splice sites and for the formation of a defined structure in the spliceosome. Thus, the proteins of the spliceosome play a decisive role in the choice of the splice site and the effeciency of splicing. 

FIGURE 18. Schematic representation of the self-splicing reaction catalyzed by group 1 introns. G is the guanosine nucleotide cofactor, P a phosphate linker... 

The study of posttranscriptional processing of RNA molecules led to one of the most exciting discoveries in modern biochemistry—the existence of RNA enzymes. The best-characterized ribozymes are the self-splicing group I introns, RNase P, and the hammerhead ribozyme (discussed below). Most of the activities of these ribozymes are based on two fundamental reactions transesterification (Fig. 26-13) and phosphodiester bond hydrolysis (cleavage). The substrate for ribozymes is often an RNA molecule, and it may even be part of the ribozyme itself. When its substrate is RNA, an RNA cat-... 

The enzymatic activity of the L-19 IVS ribozyme results from a cycle of transesterification reactions mechanistically similar to self-splicing. Each ribozyme molecule can process about 100 substrate molecules per hour and is not altered in the reaction therefore the intron acts as a catalyst. It follows Michaelis-Menten kinetics, is specific for RNA oligonucleotide substrates, and can be competitively inhibited. The kcat/Km (specificity constant) is 10s m- 1 s lower than that of many enzymes, but the ribozyme accelerates hydrolysis by a factor of 1010 relative to the uncatalyzed reaction. It makes use of substrate orientation, covalent catalysis, and metalion catalysis—strategies used by protein enzymes. 

The RNA world hypothesis requires a nucleotide polymer to reproduce itself. Can a ribozyme bring about its own synthesis in a template-directed manner The self-splicing rRNA intron of Tetrahymena (Fig. 26-26) catalyzes the reversible attack of a guanosine residue on the 5 splice junction (Fig. 26-37). If the 5 splice site and the internal guide sequence are removed from the intron, the rest of the intron can bind RNA strands paired with short oligonucleotides. Part of the remaining intact intron effectively acts as a template for the... 

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