Group II ribozymes

Figure 10.13 Phosphoryl-transfer reactions. The figure shows (a) nucleotide polymerization, (b) nucleic acid hydrolysis, (c) first cleavage of an exon-intron junction by group I ribozyme (d) and by a group II ribozyme, (e) strand transfer during transposition and (f) exon ligation during RNA splicing. (From Yang et al., 2006. Copyright 2006, with permission from Elsevier.)...

Fig. 1A-F The two-dimensional structures of various ribozymes. The ribozyme or intron portion is printed in black. The substrate or exon portion is printed in gray. Arrows indicate sites of cleavage by ribozymes A (left) the two-dimensional structure of a hammerhead ribozyme and its substrate. Outlined letters are conserved bases that are involved in catalysis right) The y-shaped structure of the hammerhead ribozyme-sub-strate complex B-F the two-dimensional structures of a hairpin ribozyme, the genomic HDV ribozyme, a group I ribozyme from Tetrahymena, a group II ribozyme from S. cer-evisiae (aiy5), and the ribozyme of RNase P from E. coli...

Group I ribozymes require au external guanosine for reactivity. Group II ribozymes do not have this requirement. They carry out catalysis via a lariat mechanism. 

The intron group I ribozymes feature common secondary structure and reaction pathways. Active sites capable of catalyzing consecutive phosphodi-ester reactions produce properly spliced and circular RNAs. Ribozymes fold into a globular conformation and have solvent-inaccessible cores as quantified by Fe(II)-EDTA-induced free-radical cleavage experiments. The Tetrahy-mem group I intron ribozyme catalyzes phosphoryl transfer between guanosine and a substrate RNA strand—the exon. This ribozyme also has been proposed to use metal ions to assist in proper folding, to activate the nucleophile, and to stabilize the transition state. ... 

Pyle AM (1996) Catalytic reaction mechanisms and structural features of group II intron ribozymes, p 75-107. In Eckstein F, Lilley DMJ (ed) Catalytic RNA, vol 10 Springer, Berlin Heidelberg New York... 

In the reactions, it is likely that Mg ions work as a Lewis acid catalyst to stabilize the leaving oxygens. This stabilization is the same as for group I II and possibly hammerhead ribozymes. But identification of the activator, encouragement of deprotonation of 2 -OH of nucleophiles, remains unclear in the reactions catalyzed by this sn RNA and group II intron ribozymes. 

There are six ribozymes that have been successfully modified and/or adapted for use in therapeutic and functional genomic applications. These are the group I introns, RNAse P, the hammerhead and hairpin motifs, the hepatitis delta ribozyme and the reverse splicing reaction of group II introns. Each of these ribozymes requires a divalent metal cation for activity (usually Mg++), which may participate in the chemistry of the cleavage/ligation reaction and/or may be important for maintaining the structure of the ribozyme. 

Single-molecule FRET has been applied to the folding of the group I intron ribozyme (Lee et al, 2007a Russell et al, 2002 Zhuang et al, 2000), the group II intron ribozyme (Steiner et al., 2008), the VS ribozyme (Pereira et al, 2008), and the interaction of a tetraloop and its receptor (Hodak et al.,... 

Group II introns are a distinct subgroup within the naturally occurring ribozymes... 

A very interesting feature of group II introns is their modularity the intronic domains can be provided separately to form a functional ribozyme (9). For example, the exD123 construct consists of a 5 -exon and Dl, D2, and D3. This construct alone is umeactive, but addition of the catalytically essential domain D5 in trans generates an active two-piece ribozyme that cleaves off the 5 -exon it is therefore a mimic for the hydrolytic pathway of the first step of splicing. Similarly, an intron construct... 

Many ribozymes and proteins that catalyze phosphoryl-transfer reactions use a mechanism employing two metal ions, and early on group II introns were hypothesized to use a similar mechanism (17). However, evidence for the existence of two metal ions in the catalytic core was only found very recently (12, 13, 18). A direct Mg + coordination of the pro-Sv oxygen of the first nucleotide of the catalytic triad is evident based on phosphorothioate substitution experiments (18). Recently, an intact group II was successfully crystallized for the first time (12). This structure confirms the metal contact from the first nucleotide of the catalytic triad and shows additional contacts to this metal ion from the second nucleotide of the catalytic triad and from the first nucleotide upstream of the bulge (Fig. 4a). This nucleotide is additionally coordinated to the second Mg + ion in the core. The distance between the two Mg + ions in the crystal stmcture is 3.9 A, which is in agreement with the proposed two-metal-ion mechanism (17). 

Group II introns naturally catalyze a broad range of reactions. The catalytic repertoire of other ribozymes was previously extended by in vitro evolution. It could be very exciting to apply this method to the versatile group II introns. 

Lehmann K, Schmidt U. Group II introns structure and catalytic versatility of large natural ribozymes. Crit. Rev. Biochem. Mol. Biol. 2003 38 249-303. 

Pyle AM. Group II introns catalysts for splicing, genomic change and evolution. In Ribozymes and RNA Catalysis. Lilley DM1, Eckstein E, eds. 2008. RCS Publishing, Cambridge, UK. 

Swisher 1, Duarte CM, Su El, Pyle AM. Visualizing the solvent-inaccessible core of a group II intron ribozyme. EMBO 1. 2001 20 2051-2061. 

Erat MC, Sigel RK. Determination of the intrinsic affinities of multiple site-specific Mg(2 -f) ions coordinated to domain 6 of a group II intron ribozyme. Inorg. Chem. 2007 46 11224-11234. 

It is an impressive list since several hundred different natural group I and group II introns have been shown to exist. In addition, many artificial active ribozymes have been prepared. 

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