Transition state stabilization by the 'high' motif of class I aminoacyl-tRNA synthetases: The case of Escherichia coli methionyl-tRNA synthetase

Methlonyl-tRNA synthetase belongs to the class I aminoacyl-tRNA synthetase family characterized both by a catalytic center built around a Rossmann Fold and by the presence of the two peptidic marker sequences HIGH and KMSKS. In this study, the role of the ₂₁HLGH₂₄ motif of Escherichia coli methlonyl-tRNA synthetase was studied In a systematic fashion by srte-dlrected mutagenesis. It is shown that the two histidine residues play a crucial role in the catalysis of the methionyl adenylate formation by participating in the stabilisation of the ATP phosphate chain during the transition state. Moreover, the results suggest the involvement of the e-imino group of histidine 21 and of the δ-imino group of histidine 24. Notably, the substitution of either the leucine or the glycine residue of the HLGH motif by alanine had no effect on the catalysis. From the data and from other studies with class I aminoacyl-tRNA synthetases, concomitant positive contributions of the HIGH and KMSKS sequences to reach the transition state of aminoacyl adenylate formation can be envisaged.