The crystal structure of the tryptic fragment of the methionyl-tRNA synthetase from Escherichia coli, complexed with ATP, has been refined to a crystallographic R-factor of 0·220, at 2·5 Å resolution (for 4433 protein atoms). In the last stages of the refinement, the simulated annealing refinement method was fully applied, contributing to a drastic improvement of the model and the identification of the missing atoms. In the final model, the root-mean-square deviation from ideality for bond distances is 0·021 Å and for angle distances is 0·054 Å. The position of the zinc ion has been confirmed and is located near the active site. The tryptic fragment is composed of two globular domains. The first domain, from the N terminus to Thr360, contains a nucleotide-binding fold into which two long polypeptides of 101 and 70 residues are inserted. The nucleotide-binding fold is strengthened by the presence of the zinc ion in the vicinity of the active site. The second domain, up to Pro526, is mainly α-helical. The C-terminal polypeptide, Phe527 to Lys551, folds back towards the first domain, making a link between the two domains. The heptapeptide 528-534 partly shapes a deep cavity that plunges into the central core of the nucleotidebinding fold, where the ATP molecule is located. The adenine ring, deeply buried in the bottom of the cleft, is blocked between the first helix HA, and the strands A and D of the β-sheet and makes no polar interaction with the enzyme. The 2′ and 3′ hydroxyl groups of the ribose, whose conformation is C2′ endo, interact with the main-chain carbonyl oxygen atoms of Ile231 and Glu241, respectively. The side-chain nitrogen atom of Lyauthor42 is at hydrogen-bonding distance from the ring oxygen O-4′ of the ribose. One of the α-phosphate oxygen atoms and one of the γ-phosphate oxygen atoms interact with the imidazole ring of Hien1, which is well conserved in many of the known synthetases; this indicates a possible crucial role for this residue in binding ATP. The β-phosphate group is linked to the mainchain carbonyl oxygen atom of Tyr15 through an intermediate water molecule. The γ-phosphate group interacts with the carbonyl oxygen atom and the side-chain of Asn17: Ly35, which is also close to the γ-phosphate (3·5 Å), is located in an equivalent position to Lyen33 of tyrosyl-tRNA synthetase, which is involved in ATP binding, and in an equivalent position to Lyen70 of glutaminyl-tRNA synthetase, which interacts with ATP phosphate groups. The length of the molecule (the extremity of the α-helical domain is located at a distance of 65 Å from the active site) is compatible to the distance between the 3′ end and the anticodon of the tRNA, rendering the peptide 456 to 476 a good candidate for an interaction with the tRNA anticodon. © 1990 Academic Press Limited.
