Structure at 2.6 A resolution of phenylalanyl-tRNA synthetase complexed with phenylalanyl-adenylate in the presence of manganese
Fishman, R.; Ankilova, V.; Moor, N.; Safro, M.
Acta Crystallographica. Section D Biological Crystallography 57(Pt 11): 1534-1544
The crystal structure of phenylalanyl-tRNA synthetase (PheRS) from Thermus thermophilus, a class II aminoacyl-tRNA synthetase, complexed with phenylalanyl-adenylate (Phe-AMP) was determined at 2.6 ANG resolution. Crystals of native PheRS were soaked in a solution containing phenylalanine and ATP in the presence of Mn2+ ions. The first step of the aminoacylation reaction proceeds within the crystals, resulting in Phe-AMP formation at the active site. Specific recognition of the phenylalanine portion of the Phe-AMP is achieved by interactions of the phenyl ring of Phe-AMP with two neighbouring residues, Phealpha258 and Phealpha260. No manganese ions were observed within the active site; their role in the formation of the transition state 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 motif 3 residues participating in the substrate binding. A sulfate ion, which was identified on the protein surface, may mediate the interactions of PheRS 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-adenylate). 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.