Proofreading in vivo: editing of homocysteine by methionyl-tRNA synthetase in the yeast Saccharomyces cerevisiae.
Homocysteine thiolactone is a product of an error-editing reaction, catalyzed by Escherichia coli methionyl-tRNA synthetase, which prevents incorporation of homocysteine into tRNA and protein, both in vitro and in vivo. Here, the thiolactone is also shown to occur in cultures of the yeast Saccharomyces cerevisiae. In yeast, the thiolactone is made from homocysteine in a reaction catalyzed by methionyl-tRNA synthetase. One molecule of homocysteine is edited as thiolactone per 500 molecules of methionine incorporated into protein. Homocysteine, added exogenously to the medium or overproduced by some yeast mutants, is detrimental to cell growth. The cost of homocysteine editing in yeast is minimized by the presence of a pathway leading from homocysteine to cysteine, which keeps intracellular homocysteine at low levels. These results not only directly demonstrate that editing of errors in amino acid selection by methionyl-tRNA synthetase operates in vivo in yeast but also establish the importance of proofreading mechanisms in a eukaryotic organism.
Jakubowski
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Department of Microbiology and Molecular Genetics, New Jersey Medical School, University of Medicine and Dentistry of New Jersey, Newark 07103.
D001345:Autoradiography; D002855:Chromatography, Thin Layer; D005838:Genotype; D006710:Homocysteine / Q000031:analogs & derivatives / Q000032:analysis / Q000378:metabolism* / Q000494:pharmacology; D007700:Kinetics; D008718:Methionine-tRNA Ligase / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D013462:Sulfur Radioisotopes
D013462:Sulfur Radioisotopes; D006710:Homocysteine; C007957:homocysteine thiolactone; D008718:Methionine-tRNA Ligase