tRNAHis guanylyltransferase adds G-1 to the 5' end of tRNAHis by recognition of the anticodon, one of several features unexpectedly shared with tRNA synthetases.
All eukaryotic tRNA(His) molecules are unique among tRNA species because they require addition of a guanine nucleotide at the -1 position by tRNA(His) guanylyltransferase, encoded in yeast by THG1. This G(-1) residue is both necessary and sufficient for aminoacylation of tRNA by histidyl-tRNA synthetase in vitro and is required for aminoacylation in vivo. Although Thg1 is presumed to be highly specific for tRNA(His) to prevent misacylation of tRNAs, the source of this specificity is unknown. We show here that Thg1 is >10,000-fold more selective for its cognate substrate tRNA(His) than for the noncognate substrate tRNA(Phe). We also demonstrate that the GUG anticodon of tRNA(His) is a crucial Thg1 identity element, since alteration of this anticodon in tRNA(His) completely abrogates Thg1 activity, and the simple introduction of this GUG anticodon to any of three noncognate tRNAs results in significant Thg1 activity. For tRNA(Phe), k(cat)/K(M) is improved by at least 200-fold. Thg1 is the only protein other than aminoacyl-tRNA synthetases that is known to use the anticodon as an identity element to discriminate among tRNA species while acting at a remote site on the tRNA, an unexpected link given the lack of any identifiable sequence similarity between these two families of proteins. Moreover, Thg1 and tRNA synthetases share two other features: They act in close proximity to one another at the top of the tRNA aminoacyl-acceptor stem, and the chemistry of their respective reactions is strikingly similar.
Jackman
Jane E
JE
Department of Biochemistry and Biophysics, University of Rochester School of Medicine, New York 14642, USA.
Phizicky
Eric M
EM
D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism*; D000926:Anticodon / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006147:Guanine / Q000378:metabolism; D007700:Kinetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009713:Nucleotidyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012354:RNA, Transfer, His / Q000737:chemistry / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D046249:Transfer RNA Aminoacylation
D000926:Anticodon; D012354:RNA, Transfer, His; D029701:Saccharomyces cerevisiae Proteins; D006147:Guanine; D009713:Nucleotidyltransferases; C504741:Thg1 protein, S cerevisiae; D000604:Amino Acyl-tRNA Synthetases