Use of a hollow fiber bioreactor for large-scale production of alpha 2-adrenoceptors in mammalian cells.
Gene cloning has revealed the existence of receptors, which are structurally similar but pharmacologically distinct. One recent example is the alpha 2-adrenergic receptor (alpha 2AR) family with three members. Preparation of membrane-embedded G-protein coupled receptor subtypes in pure form is practically impossible from natural sources and only recombinant techniques have provided possibilities to study these receptors in great detail. In this respect, both yeast and insect cell hosts have been applied successfully but no good mammalian alternative has been described for large-scale production. We describe in this report the use of S115 mouse mammary tumor cells as an effective host for large-scale production of alpha 2-adrenoceptors. These cells can be easily adapted to grow in a hollow fiber bioreactor, with up to 2.8 g of total cellular protein produced in one 0.8 m2 casette. We also show that each recombinant alpha 2-subtype exhibits their expected ligand binding properties, and suggest therefore that this system could be generally applicable to other eukaryotic plasma membrane proteins.
Ala-Uotila
S
S
Turku Centre for Biotechnology, Finland.
Marjamäki
A
A
Matikainen
M T
MT
Jalkanen
M
M
D000818:Animals; D001709:Biotechnology / Q000295:instrumentation*; D003001:Cloning, Molecular; D007700:Kinetics; D008325:Mammary Neoplasms, Experimental / Q000378:metabolism; D051379:Mice; D018341:Receptors, Adrenergic, alpha-2 / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000378:metabolism; D015016:Yohimbine / Q000378:metabolism
D018341:Receptors, Adrenergic, alpha-2; D011994:Recombinant Proteins; D015016:Yohimbine