Experimental evolution of a dense cluster of residues in tyrosyl-tRNA synthetase: quantitative effects on activity,...

A dense cluster of eight residues was identified at the crossing of two alpha-helices in tyrosyl-tRNA synthetase (TyrRS) from the thermophile Bacillus stearothermophilus. Its mechanism of evolution was characterized. Four residues of this cluster are not conserved in TyrRS from the mesophile Escherichia coli. The corresponding mutations were constructed in TyrRS(Delta1), a derivative of TyrRS from B. stearothermophilus in which the anticodon binding domain is deleted. Mutations I52L (i.e. Ile52 into Leu), M55L and L105V did not affect the activity of TyrRS(Delta1) in the pyrophosphate exchange reaction whereas T51P increased it. The kinetic stabilities of TyrRS(Delta1) and its mutant derivatives at 68.5 degreesC were determined from experiments of irreversible thermal precipitation. They were in the order L105V