Galactose-dependent expression of the recombinant Ca2(+)-binding photoprotein aequorin in yeast.
Aequorin is a Ca2(+)-binding protein that emits light upon reacting with Ca2+ and has been used as a probe for monitoring changes in the intracellular free Ca2+ concentration, [Ca2+]i. The protein consists of three components: apoaequorin (apoprotein), molecular oxygen and a chromophore. The present study was designed to conditionally express the apoaequorin cDNA of the jellyfish Aequorea victoria under the control of the GAL1 promoter in the yeast Saccharomyces cerevisiae and to investigate whether apoaequorin can be accumulated in high enough concentration in the cells to detect a Ca2+ signal in vitro. The results showed that the cells accumulated sufficient amounts of recombinant apoaequorin when incubated in the galactose-based medium and that the protein was active and not toxic to the cells, suggesting that the recombinant apoaequorin may be applicable to monitoring changes in [Ca2+]i in intact yeast cells.
Nakajima-Shimada
J
J
Division of Cell Proliferation, National Institute for Basic Biology, Okazaki, Japan.
Iida
H
H
Tsuji
F I
FI
Anraku
Y
Y
D000331:Aequorin / Q000096:biosynthesis / Q000235:genetics; D001059:Apoproteins / Q000096:biosynthesis / Q000235:genetics; D001483:Base Sequence; D002135:Calcium-Binding Proteins / Q000096:biosynthesis / Q000235:genetics*; D004247:DNA / Q000096:biosynthesis; D005690:Galactose / Q000494:pharmacology*; D015870:Gene Expression; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development
D001059:Apoproteins; D002135:Calcium-Binding Proteins; D011994:Recombinant Proteins; C060014:apoaequorin; D000331:Aequorin; D004247:DNA; D005690:Galactose