RNA methyltransferases involved in 5' cap biosynthesis

Byszewska, Magdalena; Smietanski, Miroslaw; Purta, Elzbieta; Bujnicki, Janusz M

In the SARS virus, cap1 methylation is catalyzed by a complex comprised of 2 partners: the nsp16 protein that is clearly related to the above-mentioned cap1 methyltransferases, but is inactive on its own, and a small regulatory protein nsp10 that is required for nsp16 to bind both the SAM methyl group donor and the RNA substrate. The crystal structure of the snp10-nsp16 complex showed that, in nsp16, the SAM-binding region is partially degenerated compared to "partner-independent" ribose methyltransferases, and nsp10 stabilizes the SAM binding pocket and extends the RNA-binding groove of nsp16. 43 Apart from the enzymes responsible for cap1 methylation, methyltransferases have been characterized that act on additional residues in the nascent RNA chain. Many eukaryotic organisms possess a 2 0 -O-ribose methyltransferase that methylates the 2 nd residue in mRNA and in other RNA molecules. The cap2 methyltransferase has been characterized in T. brucei (TbMTr2 44, 45 ) and in humans (CMTr2 46 ). Interestingly, while CMTr2 appears to be closely related to its human paralog CMTr1 as well as to TbMTr1, TbMTr2 is more closely related to the vaccinia virus cap1 methyltransferase. 44 In trypanosomes, a third 2 0 -O-ribose cap methyltransferase was identified and termed TbMTr3, which is responsible for the methylation of the third residue of the cap and is required for the methylation of the fourth residue. 47, 48 TbMTr3 is a close relative of TbMTr2 and of VP39, and is only remotely related to other eukaryotic cap 2 0 -O-ribose methyltransferases, which suggests that trypanosomes acquired enzymes for "additional" methylation by adapting proteins from viruses. A phylogenetic study of 2 0 -O-ribose methyltransferases revealed that the relationships between cellular and viral enzymes are quite complex, and that these proteins can vary greatly in number even in closely-related organisms. Furthermore, alveolate species were identified that possessed as many as 4 2 0 -O-ribose methyltransferases, suggesting that certain enzymes of this group may act with different substrate specificities or that new cap structures with additional methylation sites remain to be discovered. 46 Proteins with cap0 and cap1 methyltransferase activities A number of viral proteins were reported to possess both cap0 and cap1 methyltransferase activities. In most of them, this is due to the presence of multiple domains. For instance, in the human reovirus (a virus with a dsRNA genome), the cap structure formation is catalyzed by a large multidomain protein lambda 2, which in turn is a part of the reovirus core: an assembly with a relative molecular mass of 52 MDa that synthesizes, modifies and exports viral mRNA. The structure of the human reovirus core has been solved at low resolution, revealing a series of domains that include a putative guanylyltransferase domain and 2 putative methyltransferase (RFM) domains. 49 It has been suggested that the order of the domains in the lambda 2 protein corresponds to the order of the capping reactions: guanosine transfer followed by cap0 and cap1 methylation. However, comparison of domain structures suggested that the functional assignments may be different, as the RFM domain 1 shared a putative active site with the corresponding structurally characterized 2 0 -O-ribose methyltransferases, including the cap1 methyltransferase, whereas the RFM domain 2 exhibited structural similarity to the cap0 methyltransferases. 50 It should be noted th