Structure at 2.6 angstrom resolution of phenylalanyl-tRNA synthetase complexed with phenylalanyl-adenylate in the presence of manganese

The crystal structure of phenylalanyl-tRNA synthetase (PheRS) from Thermus thermophilus, a class II aminoacyl-tRNA synthetase, complexed with phenylal anyl-adenylate (Phe-AMP) was determined at 2.6 Angstrom resolution. Crystal s of native PheRS were soaked in a solution containing phenylalanine and AT P in the presence of Mn2+ ions. The first step of the aminoacylation reacti on proceeds within the crystals, resulting in Phe-AMP formation at the acti ve site. Specific recognition of the phenylalanine portion of the Phe-AMP i s achieved by interactions of the phenyl ring of Phe-AMP with two neighbour ing residues, Phe alpha 258 and Phe alpha 260. No manganese ions were obser ved within the active site; their role in the formation of the transition s tate may be assigned to a number of polar residues and water molecules. In the anomalous Fourier difference map, a divalent metal ion was detected at the interface of the alpha- and beta -subunits at a short distance from mot if 3 residues participating in the substrate binding. A sulfate ion, which was identified on the protein surface, may mediate the interactions of PheR S with DNA. Visible conformational changes were detected in the active-site area adjacent to the position of the Phe-AMP, compared with the structure of PheRS complexed with a synthetic adenylate analogue (phenylalaninyl-aden ylate). Based on the known structures of the substrate-free enzyme and its complexes with various ligands, a general scheme for the phenylalanylation mechanism is proposed.