Mechanism and substrate specificity of tRNA-guanine transglycosylases (TGTs): tRNA-modifying enzymes from the three different kingdoms of life share a common catalytic mechanism

Transfer RNA-guanine transglycosylases (TGTs) ore evolutionarily ancient enzymes, present in all kingdoms of life, catalyzing guanine exchange within their cognate tRNAs by modified 7-deazoguanine bases. Although distinct bases are incorporated into tRNA at different positions in a kingdom-specific manner the catalytic subunits of TGTs are structurally well conserved. This review provides insight into the sequential steps along the reaction pathway, substrate specificity, and conformational adaptions of the binding pockets by comparison of TGT crystal structures in complex with RNA substrates of a eubacterial and an archaebacterial species. Subs trote-binding modes indicate an evolutionarily conserved bose-exchange mechanism with a conserved aspartate serving as a nucleophile through covalent binding to C1' of the guanosine ribose moiety in an intermediate state. A second canserved aspartate seems to control the spatial rearrangement of the ribose ring along the reaction pathway and supposedly operates as a general acid/base. Water molecules inside the binding pocket accommodating interaction sites subsequently occupied by polar atoms of substrates help to elucidate subs trate-recognition and substrate-specificity features. This emphasizes the role of water molecules as general probes to map binding-site properties for structure-based drug design. Additionally, substrate-bound crystal structures allow the extraction of valuable information about the classification of the TGT superfamily into a subdivision of presumably homologous superfamilies adopting the triosephosphate isomerase type barrel fold with a standard phosphate-binding motif