Protein expression in Drosophila Schneider cells.
We expressed recombinant secreted, membrane, and cytosolic proteins in stably transfected Drosophila Schneider (SL-3) cells. To allow easy cloning of N- and C-terminal fusion proteins containing epitope- and His-tags for the detection of recombinant proteins and purification by affinity chromatography we constructed new expression vectors. To exemplify the general applicability of protein expression in Schneider cells we characterized the expression system with respect to inducibility, localization of the recombinant proteins, yields of purified proteins, and presence of posttranslational and cotranslational modifications. Secreted proteins became quantitatively N-glycosylated in SL-3 cells and the N-glycan of a Golgi-resident membrane protein was found to be Endo-H-resistant. Myristoylation of AnxXIIIb, a member of the annexin family, could be demonstrated and glycosylphosphatidylinositol-anchored proteins containing their lipid anchor were expressed efficiently in SL-3 cells. Since generation of stable cell lines and mass culture of SL-3 cells is cheap and easy, they provide an attractive eukaryotic expression system.
Benting
J
J
Cell Biology and Biophysics Programme, European Molecular Biology Laboratory, Heidelberg, D-69117, Germany.
Lecat
S
S
Zacchetti
D
D
Simons
K
K
D000469:Alkaline Phosphatase / Q000096:biosynthesis / Q000235:genetics; D000818:Animals; D017302:Annexins / Q000096:biosynthesis / Q000235:genetics; D002478:Cells, Cultured; D004330:Drosophila / Q000166:cytology*; D058851:GPI-Linked Proteins; D015870:Gene Expression; D005822:Genetic Vectors; D006031:Glycosylation; D017261:Glycosylphosphatidylinositols; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics; D019814:Myristic Acid / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis*; D014162:Transfection
D017302:Annexins; D058851:GPI-Linked Proteins; D017261:Glycosylphosphatidylinositols; D007527:Isoenzymes; D011993:Recombinant Fusion Proteins; D019814:Myristic Acid; D000469:Alkaline Phosphatase; C019839:alkaline phosphatase, placental