The expression of Escherichia coli threonine synthase and the production of threonine from homoserine in mouse 3T3 cells.
We have subcloned the coding sequence for the Escherichia coli threonine synthase gene into a eukaryotic expression vector based on the simian-virus-40 early promoter. When mouse 3T3 cells which already expressed homoserine kinase were transfected with the new plasmid, the cells were able to incorporate radioactivity from [14C]homoserine into their cell proteins. Stable cell lines were established by co-transfecting 3T3 cells with the plasmid coding for threonine synthase and another coding for homoserine kinase and G-418 (Geneticin) resistance. Cells were selected for G-418 resistance and then screened for an ability to synthesize threonine from homoserine and incorporate it into the cell protein. A cell line which expressed both the homoserine kinase and threonine synthase genes was capable of growth in a threonine-deficient medium containing homoserine.
Rowett Research Institute, Bucksburn, Aberdeen, Scotland, U.K.
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D016475:3T3 Cells / Q000201:enzymology*; D000818:Animals; D015152:Blotting, Northern; D019757:Carbon-Oxygen Lyases; D002455:Cell Division; D003001:Cloning, Molecular; D003470:Culture Media; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D006714:Homoserine / Q000378:metabolism*; D008190:Lyases / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D010770:Phosphotransferases / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor); D010957:Plasmids; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D013539:Simian virus 40 / Q000235:genetics; D013912:Threonine / Q000378:metabolism*; D014162:Transfection
D003470:Culture Media; D012333:RNA, Messenger; D013912:Threonine; D006714:Homoserine; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C021370:homoserine kinase; D008190:Lyases; D019757:Carbon-Oxygen Lyases; C023160:threonine synthase