Genetic complementation of the Saccharomyces cerevisiae leu2 gene by the Escherichia coli leuB gene.
The leucine operon of Escherichia coli was cloned on a plasmid possessing both E. coli and Saccharomyces cerevisiae replication origins. This plasmid, pEH25, transformed leuA, leuB, and leuD auxotrophs of E. coli to prototrophy; it also transformed leu2 auxotrophs of S. cerevisiae to prototrophy. beta-Isopropylmalate dehydrogenase was encoded by the leuB gene of E. coli and the leu2 gene of yeast. Verification that the leuB gene present on pEH26 was responsible for complementing yeast leu2 was obtained by isolating in E. coli several leuB mutations that resided on the plasmid. These mutant leuB- plasmids were no longer capable of complementing leu2 in S. cerevisiae. We conclude that S. cerevisiae is capable of transcribing at least a portion of the polycistronic leu operon of E. coli and can translate a functional protein from at least the second gene of this operon. The yeast Leu+ transformants obtained with pEH25, when cultured in minimal medium lacking leucine, grew with a doubling time three to four times longer than when cultured in medium supplemented with leucine.
Storms
R K
RK
Department of Biology, York University, Downsview, Ontario, Canada.
Holowachuck
E W
EW
Friesen
J D
JD
D050539:3-Isopropylmalate Dehydrogenase; D000429:Alcohol Oxidoreductases / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D005816:Genetic Complementation Test; D007930:Leucine / Q000235:genetics*; D016296:Mutagenesis; D009876:Operon; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D000429:Alcohol Oxidoreductases; D050539:3-Isopropylmalate Dehydrogenase; D007930:Leucine