Glycyl-tRNA synthetase uses a negatively charged pit for specific recognition and activation of glycine

The crystal structures of glycyl-tRNA synthetase (GlyRS) from Thermus therm ophilus, a homodimeric class II enzyme, were determined in the enzyme-subst rate and enzyme-product states corresponding to the first step of aminoacyl ation. GlyRS was cocrystallized with glycine and ATP, which were transforme d by the enzyme into glycyl-adenylate and thus gave the enzyme-product comp lex. To trap the enzyme-substrate complex, the enzyme was combined with the glycine analog ethanolamine and ATP. The ligands ape bound in fixed orient ations in the substrate-binding pocket of the N-terminal active site domain , which contains the classical class II aminoacyl-tRNA synthetase (aaRS) fo ld. Since glycine does not possess a side-chain, much of the specificity of the enzyme is directed toward excluding any additional atoms beyond the al pha-carbon atom. Several carboxylate residues of GlyRS line the glycine bin ding pocket; two of them interact directly with the alpha-ammonium group. I n addition, the enzyme utilizes the acidic character of the pro-L alpha-hyd rogen atom by contacting it via a glutamate carboxylic oxygen atom. A guani dino eta-nitrogen atom of the class II aaRS-conserved motif 2 arginine inte racts with the substrate carbonyl oxygen atom. These features serve to attr act the small amino acid substrate into the active site and to position it in the correct orientation. GlyRS uses class II-conserved residues to inter act with the ATP and the adenosine-phosphate moiety of glycyladenylate. On the basis of this similarity, we propose that GlyRS utilizes the same gener al mechanism as that employed by other class II aminoacyl-tRNA synthetases. (C) 1999 Academic Press.