Expression and functional characterization of a pHis-tagged human bradykinin B2 receptor in COS-7 cells.
A polyHis-tagged bradykinin (BK) B2 receptor (pHis-BKR) cDNA was constructed and expressed in COS-7 cells. The pHis-BKR is suitable for both immunoprecipitation and immunoblotting with anti-polyHis antibodies and can be easily purified using Ni-NTA columns. Immunochemical detection revealed a molecular mass of approximately 66 kDa. The pHis-BKR is capable of mediating BK-induced stimulation of inositol phosphate formation as well as of mitogen-activated protein kinase (MAPK) activity. Compared with the wild-type receptor (WT-BKR) the tagged receptor showed a slightly enhanced affinity towards BK but a reduced expression level. Despite these modified pharmacological properties the pHis-tagged BKR may be a useful tool for studying BKR modifications and signaling.
Müller
S
S
Institute of Biochemistry and Biophysics, Biological and Pharmaceutical Faculty, Friedrich-Schiller-University Jena, Germany.
Adomeit
A
A
Kaufmann
R
R
Appelhans
H
H
Passow
H
H
Reissmann
S
S
Liebmann
C
C
D000319:Adrenergic beta-Antagonists / Q000378:metabolism; D000345:Affinity Labels / Q000737:chemistry*; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000378:metabolism; D001665:Binding Sites; D001920:Bradykinin / Q000031:analogs & derivatives / Q000378:metabolism; D065168:Bradykinin Receptor Antagonists; D019556:COS Cells / Q000378:metabolism*; D006639:Histidine / Q000737:chemistry* / Q000276:immunology; D006801:Humans; D020128:Inhibitory Concentration 50; D007700:Kinetics; D010455:Peptides / Q000737:chemistry; D010716:Phosphatidylinositols / Q000378:metabolism; D011485:Protein Binding; D043782:Receptor, Bradykinin B2; D018002:Receptors, Bradykinin / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D014162:Transfection; D014316:Tritium
D000319:Adrenergic beta-Antagonists; D000345:Affinity Labels; D000894:Anti-Inflammatory Agents, Non-Steroidal; D065168:Bradykinin Receptor Antagonists; D010455:Peptides; D010716:Phosphatidylinositols; D043782:Receptor, Bradykinin B2; D018002:Receptors, Bradykinin; D014316:Tritium; D006639:Histidine; C065679:icatibant; D001920:Bradykinin