title: Non-essential role of lysine residues for the catalytic activities of aspartyl-tRNA synthetase and comparison with other aminoacyl-tRNA synthetases.
abstract: Essential lysine residues were sought in the catalytic site of baker's yeast aspartyl-tRNA synthetase (an alpha 2 dimer of Mr 125,000) using affinity labeling methods and periodate-oxidized adenosine, ATP, and tRNA(Asp). It is shown that the number of periodate-oxidized derivatives which can be bound to the synthetase via Schiff's base formation with epsilon-NH2 groups of lysine residues exceeds the stoichiometry of specific substrate binding. Furthermore, it is found that the enzymatic activities are not completely abolished, even for high incorporation levels of the modified substrates. The tRNA(Asp) aminoacylation reaction is more sensitive to labeling than is the ATP-PPi exchange one; for enzyme preparations modified with oxidized adenosine or ATP this activity remains unaltered. These results demonstrate the absence of a specific lysine residue directly involved in the catalytic activities of yeast aspartyl-tRNA synthetase. Comparative labeling experiments with oxidized ATP were run with several other aminoacyl-tRNA synthetases. Residual ATP-PPi exchange and tRNA aminoacylation activities measured in each case on the modified synthetases reveal different behaviors of these enzymes when compared to that of aspartyl-tRNA synthetase. When tested under identical experimental conditions, pure isoleucyl-, methionyl-, threonyl- and valyl-tRNA synthetases from E. coli can be completely inactivated for their catalytic activities; for E. coli alanyl-tRNA synthetase only the tRNA charging activity is affected, whereas yeast valyl-tRNA synthetase is only partly inactivated. The structural significance of these experiments and the occurrence of essential lysine residues in aminoacyl-tRNA synthetases are discussed.(ABSTRACT TRUNCATED AT 250 WORDS)
mesh_terms: D000241:Adenosine / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000345:Affinity Labels; D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism*; D001226:Aspartate-tRNA Ligase / Q000378:metabolism*; D002384:Catalysis; D008239:Lysine / Q000502:physiology*; D010084:Oxidation-Reduction; D010449:Peptide Mapping; D012349:RNA, Transfer, Asp / Q000494:pharmacology
chemical_list: D000345:Affinity Labels; D012349:RNA, Transfer, Asp; D000255:Adenosine Triphosphate; D000604:Amino Acyl-tRNA Synthetases; D001226:Aspartate-tRNA Ligase; D008239:Lysine; D000241:Adenosine