The GIR1 branching ribozyme is a 179 nt ribozyme with a structural resemblance to a group I ribozyme.[2] It is found within a complex type of group I introns also termed twin-ribozyme introns.[3] Rather than splicing, it catalyses a branching reaction in which the 2'OH of an internal residue is involved in a nucleophilic attack at a nearby phosphodiester bond.[4] As a result, the RNA is cleaved at an internal processing site (IPS), leaving a 3'OH and a downstream product with a tiny lariat at its 5' end. The lariat has the first and the third nucleotide joined by a 2',5' phosphodiester bond and is referred to as 'the lariat cap' because it caps an intron-encoded mRNA. The resulting lariat cap seems to contribute by increasing the half-life of the HE mRNA,[4][5] thus conferring an evolutionary advantage to the HE.
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The GIR1 ribozyme was originally discovered during the functional characterization of the introns from the extrachromosomal rDNA of the Didymium iridis protist. A combination of deletion and in vitro self-splicing analyses revealed a twin-ribozyme intron organization: two distinct ribozyme domains within the intron.[3]
The twin-ribozyme introns represent some of the most complex organized group I introns known and consist of a homing endonuclease gene (HEG: I-DirI homing endonuclease) embedded in two functionally distinct catalytic RNA domains. One of the catalytic RNAs is a conventional group I intron ribozyme (GIR2) responsible for the intron splicing and reverse splicing, as well as intron RNA circularization. The other catalytic RNA domain is the group I-like ribozyme (GIR1) directly involved in homing endonuclease mRNA maturation.
| GIR1 Branching Ribozyme | |
|---|---|
| Conserved secondary structure of GIR1 | |
| Identifiers | |
| Symbol | GIR1 |
| Rfam | RF01807 |
| Other data | |
| RNA type | Intron |
| Domain(s) | Naegleria |
In vitro, DiGIR1 catalyses three different reactions. The first one consists in hydrolysis of the scissil phosphate at the IPS site. This is the cleavage reaction observed with the full-length intron and several length variants with a relative low rate. The hydrolytic cleavage is irreversible and is considered as an in vitro artefact resulting from a miss-folding of the catalytic site to present the branch nucleotide (BP) correctly for the reaction. The second reaction,the natural one, is the branching reaction, in which a transesterification at the IPS site results in the cleavage of the RNA with a 3'OH and a downstream lariat cap made by joining of the first and the third nucleotide by a 2'-5' phosphodiester bond.[4]
These products are the only products observed by analysis of cellular RNA.[5][6] This branching reaction is in equilibrium with a third one: a ligation reaction. It is a very efficient reaction and it tends to mask the branching reaction during the in vitro branching experiments with the full-length intron and length variants that include more than 166 nucleotides upstream of the IPS.
GIR1 models have been created using biochemical and mutational data.[1] The structure contains an extended substrate domain which contains a GoU pair. The pair differs from the typical group 1 ribozyme nucleophilic residue, the J8/7 region has been reduced.[1] These findings provide the basis for an evolutionary mechanism that accounts for the change from group I splicing ribozyme to the branching GIR1 architecture. This mechanism could potentially be applied to other large RNAs such as the ribonuclease P.[1]