Recent analyses identified a putative catalytic tetrad K-D-K-E common to several families of site-specific methyltransferases (MTases) that modify 2'-hydroxyl groups of ribose in mRNA, rRNA and tRNA (designated the RrmJ class after one of the structurally characterized members; 1eiz in Protein Data Bank) [Genome Biol. 2(9) (2001) 38]. Subsequently, three residues of the tetrad (K-D-K) were shown to be essential for catalysis in RrmJ [J. Biol. Chem. 277 (2002) 41978]. Here, we report identification of a similar conserved tetrad (K-D-K-H) in the family of snoRNA-guided ribose 2'-O-MTases related to fibrillarin (represented by the Mj0697 protein structure; 1fbn in PDB). The corresponding functional groups of putative catalytic tetrads of RrmJ and Mj0697 may be superimposed in space. However, one of the invariant residues (K(164) in RrmJ and K(179) in Mj0697) is observed in two distinct locations in the primary sequence, suggesting an interesting case of 'migration' of the conserved side chain within the framework of the active site. RrmJ and Mj0697 sequences were used as starting points to carry out comprehensive sequence database searches, resulting in identification of a similar conserved tetrad (and hence, prediction of a ribose 2'-O-specificity) in several families of putative MTases, including TlyA hemolysins, novel proteins from Trypanosoma, and large multidomain proteins from Flaviviriruses, Nidoviruses, and Alphaviruses. The results of our analysis of phylogenetic relationships in the RrmJ/fibrillarin superfamily provide insight into the evolution of site-specific and snoRNA-guided ribose 2'-O-MTases from a common ancestor.