Photoaffinity labelling of atrial natriuretic factor (ANF)-R1 receptor by underivatized 125I-ANF. Involvement of lipid peroxidation.
In bovine adrenal zona glomerulosa, atrial natriuretic factor (ANF) exerts its physiological effect through high-affinity binding to specific membrane receptors. On studying further the molecular properties of the ANF receptor binding domain, we have observed that incubation of intact or solubilized bovine adrenal zona glomerulosa membranes with 125I-ANF-(99-126) followed by u.v. irradiation results in the irreversible labelling of a 130 kDa protein corresponding to the ANF-RI receptor. This process is time-, protein- and 125I-ANF-dependent. The apparently covalent nature of this complex is documented by its resistance to heat, guanidine hydrochloride, urea and trichloroacetic acid denaturation. Photolabelling with underivatized 125I-ANF is much more efficient with the ANF-R1 than with the ANF-R2 receptor. After photolysis, the covalently linked 125I-ANF is still sensitive to digestion by carboxypeptidase A, suggesting that ANF is linked by its N-terminal end to the receptor upon u.v. irradiation and that its C-terminal end is still freely accessible. Aerobic conditions and lipids are required for the photolabelling, suggesting a role in this process for malondialdehyde, a highly reactive secondary product associated with u.v.-induced lipid peroxidation. This simple method should provide a powerful tool in the accurate characterization of the hormone-binding domain of the ANF receptor.
Larose
L
L
Laboratory of Molecular Pharmacology, Clinical Research Institute of Montreal, Canada.
McNicoll
N
N
Rondeau
J J
JJ
Escher
E
E
De Lean
A
A
D000345:Affinity Labels / Q000378:metabolism*; D000818:Animals; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D007457:Iodine Radioisotopes; D007700:Kinetics; D015227:Lipid Peroxidation; D008970:Molecular Weight; D017461:Receptors, Atrial Natriuretic Factor; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism*; D015384:Zona Glomerulosa / Q000378:metabolism*
D000345:Affinity Labels; D007457:Iodine Radioisotopes; D011956:Receptors, Cell Surface; D009320:Atrial Natriuretic Factor; D017461:Receptors, Atrial Natriuretic Factor