Mutational separation of two pathways for editing by a class I tRNA synthetase.
Aminoacyl tRNA synthetases (aaRSs) catalyze the first step in protein biosynthesis, establishing a connection between codons and amino acids. To maintain accuracy, aaRSs have evolved a second active site that eliminates noncognate amino acids. Isoleucyl tRNA synthetase edits valine by two tRNA(Ile)-dependent pathways: hydrolysis of valyl adenylate (Val-AMP, pretransfer editing) and hydrolysis of mischarged Val-tRNA(Ile) (posttransfer editing). Not understood is how a single editing site processes two distinct substrates--an adenylate and an aminoacyl tRNA ester. We report here distinct mutations within the center for editing that alter adenylate but not aminoacyl ester hydrolysis, and vice versa. These results are consistent with a molecular model that shows that the single editing active site contains two valyl binding pockets, one specific for each substrate.
Hendrickson
Tamara L
TL
The Scripps Research Institute and, The Skaggs Institute for Chemical Biology, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Nomanbhoy
Tyzoon K
TK
de Crécy-Lagard
Valérie
V
Fukai
Shuya
S
Nureki
Osamu
O
Yokoyama
Shigeyuki
S
Schimmel
Paul
P
D000249:Adenosine Monophosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D004951:Esterification; D006868:Hydrolysis; D007532:Isoleucine / Q000737:chemistry; D007533:Isoleucine-tRNA Ligase / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D014176:Protein Biosynthesis / Q000502:physiology*; D011487:Protein Conformation; D012346:RNA, Transfer, Amino Acyl / Q000096:biosynthesis; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000502:physiology; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D014633:Valine / Q000737:chemistry
D029968:Escherichia coli Proteins; D012346:RNA, Transfer, Amino Acyl; D011993:Recombinant Fusion Proteins; D007532:Isoleucine; D000249:Adenosine Monophosphate; D007533:Isoleucine-tRNA Ligase; D014633:Valine