High human GLUT1, GLUT2, and GLUT3 expression in Schizosaccharomyces pombe.
In this study, three subfamily members of the human 12-transmembrane-domain cell-surface receptors GLUT1, GLUT2, and GLUT3 were heterologously expressed in the fission yeast Schizosaccharomyces pombe utilizing GST-GLUT fusion proteins. These fusion proteins were driven by the full-length nmt1 promoter (Pnmt1) derived from S. pombe. The transcription levels of the GST-GLUT fusion proteins were very high upon induction by removing thiamine from the media. One-step purification of the recombinant fusion proteins was achieved by GST-affinity chromatography. Approximately 300 microg of highly purified fusion protein were obtained from 3 g of wet cell paste (1 liter of cell culture), indicating that human membrane proteins can be efficiently expressed and purified in the fission yeast. With its available extensive genetic information and ease of genetic manipulation, the fission yeast is potentially a highly efficient host to express eukaryotic membrane proteins.
Yang
Yuxin
Y
College of Bioengineering, Chongqing University, Shapingba, Chongqing, 400044, China.
Hu
Zongli
Z
Liu
Zhizhao
Z
Wang
Yi
Y
Chen
Xuqing
X
Chen
Guoping
G
D015153:Blotting, Western; D051272:Glucose Transporter Type 1 / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D051273:Glucose Transporter Type 2 / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D051274:Glucose Transporter Type 3 / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D005982:Glutathione Transferase / Q000096:biosynthesis / Q000302:isolation & purification; D006801:Humans; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D012568:Schizosaccharomyces / Q000235:genetics / Q000378:metabolism*
D051272:Glucose Transporter Type 1; D051273:Glucose Transporter Type 2; D051274:Glucose Transporter Type 3; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase