Aminoacyl transfer rate dictates choice of editing pathway in threonyl-tRNA synthetase.
Aminoacyl-tRNA synthetases hydrolyze aminoacyl adenylates and aminoacyl-tRNAs formed from near-cognate amino acids, thereby increasing translational fidelity. The contributions of pre- and post-transfer editing pathways to the fidelity of Escherichia coli threonyl-tRNA synthetase (ThrRS) were investigated by rapid kinetics. In the pre-steady state, asymmetric activation of cognate threonine and noncognate serine was observed in the active sites of dimeric ThrRS, with similar rates of activation. In the absence of tRNA, seryl-adenylate was hydrolyzed 29-fold faster by the ThrRS catalytic domain than threonyl-adenylate. The rate of seryl transfer to cognate tRNA was only 2-fold slower than threonine. Experiments comparing the rate of ATP consumption to the rate of aminoacyl-tRNA(AA) formation demonstrated that pre-transfer hydrolysis contributes to proofreading only when the rate of transfer is slowed significantly. Thus, the relative contributions of pre- and post-transfer editing in ThrRS are subject to modulation by the rate of aminoacyl transfer.
Minajigi
Anand
A
Cell and Molecular Biology Program, University of Vermont, Burlington, VT 05405, USA.
Francklyn
Christopher S
CS
D000255:Adenosine Triphosphate / Q000737:chemistry; D001665:Binding Sites; D020134:Catalytic Domain; D004926:Escherichia coli / Q000201:enzymology*; D006868:Hydrolysis; D007700:Kinetics; D008954:Models, Biological; D008956:Models, Chemical; D008957:Models, Genetic; D017393:RNA Editing; D012346:RNA, Transfer, Amino Acyl / Q000737:chemistry*; D012694:Serine / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D013912:Threonine / Q000737:chemistry
D012346:RNA, Transfer, Amino Acyl; D012997:Solvents; D013912:Threonine; D012694:Serine; D000255:Adenosine Triphosphate