Complementation of a pgk deletion mutation in Saccharomyces cerevisiae with expression of the phosphoglycerate-kinase gene from the hyperthermophilic Archaeon Sulfolobus solfataricus.
The gene encoding phosphoglycerate kinase (PGK) from the Archaeon Sulfolobus solfataricus, an organism growing optimally at 87 degrees C, was inserted into a yeast expression vector under the control of the galactose-inducible GAL1 yeast promoter. This vector was then transformed into a pgk::TRP1 yeast mutant, a strain inhibited for growth on galactose or glucose due to its lack of PGK enzyme. Slow-growing transformants were obtained on galactose plates at 37 degrees C, but not 28 degrees C. These transformants contained low levels of transcripts of the heterologous gene and low amounts of thermostable PGK activity. Weak expression of the hyperthermophile gene in yeast, a mesophile, therefore enabled complementation of the yeast pgk defect at 37 degrees C but not at 28 degrees C.
Piper
P W
PW
Department of Biochemistry and Molecular Biology, University College London, London WC1E 6BT, UK.
Emson
C
C
Jones
C E
CE
Cowan
D A
DA
Fleming
T M
TM
Littlechild
J A
JA
D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004795:Enzyme Stability; D017353:Gene Deletion; D015870:Gene Expression; D005798:Genes, Bacterial; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D010735:Phosphoglycerate Kinase / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics*; D016944:Sulfolobus / Q000201:enzymology* / Q000235:genetics*; D014170:Transformation, Genetic
D017931:DNA Primers; D010735:Phosphoglycerate Kinase