Further development of the locus control region/murine erythroleukemia expression system: high level expression and characterization of recombinant human calcitonin receptor.
In order to demonstrate the potential of the LCR/MEL expression system for the expression of seven-transmembrane helix receptors the human calcitonin receptor (hCTR) has been cloned and expressed at high levels in this system. Using the newly developed single-step expression vectors pEV and pNV, stable recombinant MEL cells which express the hCTR at approximately 6 pmol of receptor/mg of total cell protein (60 pmol/mg of membrane protein), or 1.9 x 10(6) receptors per cell, have been generated. Ligand binding studies have shown that the cloned receptor expressed in MEL cells has properties which are indistinguishable from those of the native receptor in T47D cells. It is suggested that MEL cells expressing seven-transmembrane helix receptors provide an efficient and readily attainable source of material for ligand binding experiments using these receptors.
Needham
M
M
Vascular, Inflammatory & Musculo-skeletal Research, ZENEXA Pharmaceuticals, Macclesfield, Cheshire, United Kingdom.
Egerton
M
M
Millest
A
A
Evans
S
S
Popplewell
M
M
Cerillo
G
G
McPheat
J
J
Monk
A
A
Jack
A
A
Johnstone
D
D
D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D002116:Calcitonin / Q000378:metabolism; D003001:Cloning, Molecular; D016384:Consensus Sequence; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute; D051379:Mice; D008969:Molecular Sequence Data; D010442:Peptide Chain Initiation, Translational; D018003:Receptors, Calcitonin / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D018003:Receptors, Calcitonin; D011993:Recombinant Fusion Proteins; D002116:Calcitonin