Transfer RNA-dependent translocation of misactivated amino acids to prevent errors in protein synthesis.
Misactivation of amino acids by aminoacyl-tRNA synthetases can lead to significant errors in protein synthesis that are prevented by editing reactions. As an example, discrete sites in isoleucyl-tRNA synthetase for amino acid activation and editing are about 25 A apart. The details of how misactivated valine is translocated from one site to the other are unknown. Here, we present a kinetic study in which a fluorescent probe is used to monitor translocation of misactivated valine from the active site to the editing site. Isoleucine-specific tRNA, and not other tRNAs, is essential for translocation of misactivated valine. Misactivation and translocation occur on the same enzyme molecule, with translocation being rate limiting for editing. These results illustrate a remarkable capacity for a specific tRNA to enhance amino acid fine structure recognition by triggering a unimolecular translocation event.
Nomanbhoy
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Skaggs Institute for Chemical Biology, Scripps Research Institute, Beckman Center, La Jolla, California 92037, USA.
Hendrickson
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Schimmel
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D000255:Adenosine Triphosphate / Q000378:metabolism; D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism; D001665:Binding Sites; D003838:Deoxyadenine Nucleotides / Q000378:metabolism; D004926:Escherichia coli; D005456:Fluorescent Dyes; D007700:Kinetics; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics; D012343:RNA, Transfer / Q000235:genetics*; D012355:RNA, Transfer, Ile / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics; D013050:Spectrometry, Fluorescence; D046249:Transfer RNA Aminoacylation / Q000235:genetics*; D014633:Valine / Q000378:metabolism
D003838:Deoxyadenine Nucleotides; D005456:Fluorescent Dyes; D011506:Proteins; D012355:RNA, Transfer, Ile; D016601:RNA-Binding Proteins; D000255:Adenosine Triphosphate; D012343:RNA, Transfer; D000604:Amino Acyl-tRNA Synthetases; D014633:Valine; C026600:2'-deoxyadenosine triphosphate