The hsp150Δ-carrier confers secretion competence to the rat nerve growth factor receptor ectodomain in Saccharomyces cerevisiae
✍ Scribed by Simonen, Marjo; Vihinen, Helena; Jämsä, Eija; Arumäe, Urmas; Kalkkinen, Nisse; Makarow, Marja
- Book ID
- 102648026
- Publisher
- John Wiley and Sons
- Year
- 1996
- Tongue
- English
- Weight
- 798 KB
- Volume
- 12
- Category
- Article
- ISSN
- 0749-503X
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✦ Synopsis
When the extracellular domain of rat low-affinity nerve growth factor receptor (NGFR,) was synthesized in Saccharomyces cerevisiae with the signal peptide of invertase, NGFR, was translocated to the endoplasmic reticulum (ER) and retained there. However, when NGFR, was fused to the C-terminus of the hsp150A-carrier, the hsp150A-NGFRe fusion protein was efficiently secreted to the growth medium with no apparent retention in the ER. The NGFR, portion was disulphide-bonded and its single N-glycosylation site was occupied. The hspl50Acarrier is an N-terminal signal peptide-containing fragment of a yeast secretory glycoprotein. Hsp 150A-NGFRe, harvested from the culture medium, inhibited the cross-linking of ['251]NGF to authentic NGFR on the surface of human melanoma cells. Moreover, [12'I]NGF could be chemically cross-linked to secretory hsp150A-NGFRe, suggesting that the NGFR, portion had adopted a ligand-binding conformation. However, inhibition of the cross-linking by unlabelled NGF was less effective than in the case of the authentic receptor. The hspl50A-carrier may have potential in the production of mammalian proteins, which require elaborate folding and disulphide formation in the ER.