THE FIRST STEP OF AMINOACYLATION AT THE ATOMIC-LEVEL IN HISTIDYL-TRANSFER-RNA SYNTHETASE

The crystal structure of an enzyme-substrate complex with histidyl-tRN A synthetase from Escherichia coli, ATP, and the amino acid analog his tidinol is described and compared with the previously obtained enzyme- product complex with histidyl-adenylate. An active site arginine, Arg- 259, unique to all histidyl-tRNA synthetases, plays the role of the ca talytic magnesium ion seen in seryl-tRNA synthetase. When Arg-259 is s ubstituted with histidine, the apparent second order rate constant (k( cat)/K-m) for the pyrophosphate exchange reaction and the aminoacylati on reaction decreases 1,000-fold and 500-fold, respectively. Crystals soaked with MnCl2 reveal the existence of two metal binding sites betw een beta- and gamma-phosphates; these sites appear to stabilize the co nformation of the pyrophosphate. The use of both conserved metal ions and arginine in phosphoryl transfer provides evidence of significant e arly functional divergence of class II aminoacyl-tRNA synthetases.