Purification and characterisation of B2 bradykinin receptor from rat uterus.
A B2 bradykinin (BK) receptor was solubilised and partially purified from rat uterine membranes by a combination of ammonium sulphate precipitation, desalting on Sephadex G-50, and hydroxyapatite and wheat germ agglutinin affinity chromatography. The partially purified BK receptor, enriched 1,500-fold, was then cross-linked to 125I-Tyr0-BK using disuccinimidyl suberate and purified to homogeneity as a single protein species on two-dimensional gel electrophoresis with a molecular mass of 81 kDa. This molecular size was in agreement with the value of 80-120 kDa estimated from Sephacryl 300 size exclusion column chromatography of the B2 receptor. The partially purified and the crude solubilised B2 BK receptor from rat uterus showed similar affinities for BK and the BK analogues iodo-Tyr0-Bk, D-Phe7-BK, and des-Arg9-BK, indicating that the ligand binding specificity of the receptor had been retained during the purification procedures. The biochemical properties of the solubilised B2 BK receptor correspond to those of a hydrophobic acidic glycoprotein (isoelectric focusing gave a value of 4.5-4.7) that binds specifically to wheat germ agglutinin but has no affinity for either concanavalin A or lentil lectin, suggesting the absence of terminal mannose or glucose residues.
Yaqoob
M
M
Department of Biochemistry, Sandoz Institute for Medical Research, London, England.
Snell
C R
CR
D000818:Animals; D001920:Bradykinin / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002845:Chromatography; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D017886:Durapatite; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D007700:Kinetics; D008970:Molecular Weight; D051381:Rats; D017207:Rats, Sprague-Dawley; D018002:Receptors, Bradykinin / Q000302:isolation & purification* / Q000378:metabolism*; D014599:Uterus / Q000378:metabolism*
D018002:Receptors, Bradykinin; D017886:Durapatite; D001920:Bradykinin