Characterization of the functional domains of the natriuretic peptide receptor/guanylate cyclase by radiation inactivation.
Radiation inactivation has been used to evaluate the molecular size of domains responsible for atrial natriuretic peptide (ANP)-binding and cyclase functions of the ANP receptor/guanylate cyclase. Two types of inactivation curves were observed for cyclase function in both adrenal cortex and aortic smooth muscle cells: 1) biphasic with enhanced guanylate cyclase activity after exposure to low radiation doses and 2) linear after preincubation of membrane proteins with 0.5 microM ANP or solubilization with Triton X-100. The existence of an inhibitory component was the simplest model that best explained the types of radiation curves obtained. Activation of guanylate cyclase by ANP or Triton X-100 could occur via the dissociation of this inhibitory component from the catalytic domain. On the other hand, the loss of ANP-binding activity was linear with increasing radiation exposures under basal, ANP treatment, and Triton X-100 solubilization conditions. Radiation inactivation sizes of about 30 kDa for cyclase function, 20 kDa for ANP-binding function, and 90 kDa for inhibitory function were calculated. These studies suggest that the ANP receptor/guanylate cyclase behaves as a multidomain protein. The results obtained by radiation inactivation of the various biological functions of this receptor are compatible with the hypothesis of an intramolecular inhibitory domain repressing the guanylate cyclase catalytic domain within its membrane environment.
Research Centre of Hôtel-Dieu Hospital, University of Montreal, Quebec, Canada.
J
J
Tremblay
C
C
Huot
C
C
Koch
M
M
Potier
D000311:Adrenal Glands / Q000201:enzymology / Q000528:radiation effects; D000818:Animals; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D002478:Cells, Cultured; D003902:Detergents; D004307:Dose-Response Relationship, Radiation; D004789:Enzyme Activation; D005720:Gamma Rays; D006162:Guanylate Cyclase / Q000378:metabolism* / Q000528:radiation effects; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism* / Q000528:radiation effects; D009131:Muscle, Smooth, Vascular / Q000201:enzymology / Q000528:radiation effects; D017830:Octoxynol; D011092:Polyethylene Glycols; D051381:Rats; D011921:Rats, Inbred WKY; D017461:Receptors, Atrial Natriuretic Factor; D011956:Receptors, Cell Surface / Q000378:metabolism*; D012995:Solubility
D003902:Detergents; D008565:Membrane Proteins; D011956:Receptors, Cell Surface; D011092:Polyethylene Glycols; D009320:Atrial Natriuretic Factor; D017830:Octoxynol; D006162:Guanylate Cyclase; D017461:Receptors, Atrial Natriuretic Factor