CO-SOLUBILIZATION OF BRADYKININ B-2 RECEPTORS AND ANGIOTENSIN-CONVERTING ENZYME FROM GUINEA-PIG LUNG MEMBRANES

Bradykinin B-2 receptor-like binding activity was solubilized from gui nea pig lung using the zwitterionic detergent 3-[(3-cholamidopropyl)di methylammonio]-1-propa (Chaps). The binding of [H-3]bradykinin to the soluble fraction was time-dependent and saturable. Scatchard analysis of equilibrium binding data indicated that the soluble extract contain ed a single class of binding sites with a K-d Of 696 pM and a B-max of 57 fmol/mg protein. Unlabelled bradykinin and B-2 antagonists inhibit ed the binding of [H-3]bradykinin to Chaps-solubilized extracts with r elative potencies similar to those observed with the low-affinity memb rane-bound binding sites. Following partial purification of the solubl e preparation, using anion exchange (DEAE-Sephacel) and gel filtration (Aca 34) column chromatography steps, two peaks eluted off the column were able to bind [H-3]bradykinin and have molecular masses of 168 an d 98.5 kDa. The former seems to represent binding of bradykinin to ang iotensin converting enzyme (ACE, EC 3.4.15.1) and the latter binding t o bradykinin receptor. Using purified commercial ACE, we show that the binding of [H-3]bradykinin to ACE can easily be distinguished from th at of the bradykinin receptor, since both B-1 and B-2 ligands were abl e to inhibit bradykinin binding with affinities clearly different from that expected for a bradykinin receptor.