kallidin and Neuroblastoma

The peptides bradykinin and kallidin are released in response to noxious stimuli and mediate various physiological effects, including a direct stimulation of nociceptive afferent neurones. The nature of the receptor molecules through which these ligands act is presently unknown. We synthesised an iodinatable photoaffinity probe, N epsilon-4-azidosalicylylkallidin, and used it in an attempt to identify candidate bradykinin receptors on the NG108-15 neuroblastoma X glioma hybrid cell line. The ligand bound in subdued light to a particulate fraction of NG108-15 tumours and could be displaced by bradykinin with an IC50 of 0.33 nM. In a physiological assay, it behaved as an agonist equipotent with bradykinin. Gel analysis of the labelled products after photolysis of the iodinated ligand in the presence of NG108-15 cells or tumour membranes revealed bradykinin-blockable labelling of a glycoprotein with an Mr of 166,000. The probe was also able to label purified commercial angiotensin converting enzyme. The band labelled in NG108-15 cells was immunoprecipitable with a polyclonal antiserum to angiotensin converting enzyme, an enzyme shown to be present in low amounts in these preparations by direct binding using the iodinatable specific ligand MK351A.

Topics: Affinity Labels; Angiotensin-Converting Enzyme Inhibitors; Azides; Bradykinin; Calcium; Dipeptides; Electrophoresis, Polyacrylamide Gel; Glioma; Hybrid Cells; Immunosorbent Techniques; Iodine Radioisotopes; Kallidin; Molecular Structure; Molecular Weight; Neuroblastoma; Peptide Hydrolases; Peptidyl-Dipeptidase A; Photolysis; Receptors, Bradykinin; Receptors, Neurotransmitter; Tumor Cells, Cultured

Topics: Affinity Labels; Amino Acid Sequence; Animals; Azides; Binding, Competitive; Bradykinin; Cell Line; Glioma; Hybrid Cells; Kallidin; Kinetics; Molecular Sequence Data; Neuroblastoma; Peptidyl-Dipeptidase A; Receptors, Bradykinin; Receptors, Neurotransmitter