Spliceosome splicing reaction

Fig. 12 The spliceosome splicing reaction. In the first step, the 2 -OH of an adenosine residue that is conserved in the intron attacks the phosphorus at the 5 splice site and generates an intron-3 -exon 2 intermediate and a free 5 exon 1. In the second step, the free 3 -OH of the 5 exon attacks the phosphorus at the 3 splice site to produce ligated exons and an excised intron...

Of the five snRNAs, U2 and U6 interact with the reaction site (the 5 splice site and the branch point) in the first chemical step. These two snRNAs are known to anneal together to form a stable-based paired structure in the absence of proteins and in the presence of ions as shown in Fig. 13, with U2 acting as an inducer molecule that displaces the U4 (that is an antisense molecule that regulates the catalytic function of U6 RNA) from the initially formed U4-U6 duplex. The secondary (or higher ordered) structure of the U2-U6 complex consists of the active site of the spliceosome. Recent data suggests that these two snRNAs function as the catalytic domain of the spliceosome that catalyzes the first step of the splicing reaction [145]. 

The known catalytic repertoire of ribozymes continues to expand. Some virusoids, small RNAs associated with plant RNA viruses, include a structure that promotes a self-cleavage reaction the hammerhead ribozyme illustrated in Figure 26-25 is in this class, catalyzing the hydrolysis of an internal phosphodiester bond. The splicing reaction that occurs in a spliceosome seems to rely on a catalytic center formed by the U2, U5, and U6 snRNAs (Fig. 26-16). And perhaps most important, an RNA component of ribosomes catalyzes the synthesis of proteins (Chapter 27). 

The mechanism of self-splicing in this case is somewhat different from that observed in the spliceosome reaction (fig. 28.21). First the 3 hydroxyl group of the guanosine cofactor attacks the phosphodiester bond at the 5 splice site. This is followed by another transesterification reaction in which the 3 hydroxyl group of the upstream RNA attacks the phosphodiester bond at the 3 splice site, thereby completing the splicing reaction. The final reaction products include the spliced rRNA and the excised oligonucleotide. 

Since its initial discovery, self-splicing has been found to occur for RNAs from a wide variety of organisms. Certain precursor RNAs that exhibit self-splicing produce lariats, just like those seen in the commonly observed splicing reactions that are catalyzed by spliceosomes (see fig. 28.19). These findings suggest that at one time all splicing reactions were RNA-catalyzed. 

The small nuclear ribonucleoprotein particles (snRNPs or snurps ) that carry out the splicing reaction use RNA-RNA basepairing to select the splice sites. Almost all intron-exon junctions contain the sequence AG-GU with the GU beginning the intron sequence. Furthermore, the consensus sequence for the beginning of the intron has a longer sequence complementary to the U1 RNA. Thus, assembly of the splicing complex, called the spliceosome, starts when the RNA component of the U1 snRNP base pairs with the junction between the 3 end of the exon and the 5 end of the intron. See Figure 12-13. 

Spliceosome assembly pathway showing the conserved GU-AG residues at the intron boundaries and the adenylate residue at the lariat branch site. Note that the dissociation of the U4U6 complex after the first transesterification step leads to the formation of the U2U6 complex which is critical to completion of the splicing reaction. 

Figure 11.23 The fomation of a lariat in group II intron splicing The 5 -phosphate at the scissile bond of Nj is transferred to 2 -hydroxyl of N, (the lariat A residue) to form a lariat structure in the initial step of splicing reaction mediated by group II intron and spliceosome.

The non-self-splicing reaction of pre-mRNAs takes place within the dynamic environment of spliceosome (122). The spliceosome is a multicomponent complex containing pre-mRNA, U-series snRNAs complexed with proteins (called snRNPs), and non-snRNP splicing factors. The major nucleoplasmic snRNAs (Ul, U2, U4, U5, and U6) range in size from 106 nt (U6) to 189 nt (U2), have... 

The removal of introns from pre-messenger RNAs in eukaryotes is catalyzed by the spliceosome, which is a large ribonucleoprotein consisting of at least 70 proteins and five small nuclear RNAs (snRNA) [144]. This splicing pathway involves two phosphotransfer reactions. In the first step, the 5 splice site is attacked by a 2 hydroxy group of an adenosine nucleotide within the intron [indicated by A in Fig. 12] that corresponds to the branch point in the lariat intermediate (Fig. 12,middle). In the second step, the 3 -OH group of the free 5 exon attacks the phosphodiester bond between the intron and... 

Figure 11-4. Splicing of a eukaryotic RNA transcript. A hypothetical hnRNA with two exons (EI and E2) and a single, large intron (I) is shown. Splicing can be divided into two main reactions initial attack of ribose near an A residue within the intron on the splice donor followed by attack of the newly available 3 end of exon I (EI) on the 5 end of exon 2 (E2) with coincident release of the intron. Special sequences surround the splice donor and acceptor sites. All steps occur within the spliceosome complex.

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