Divalent cations regulate glucagon binding. Evidence for actions on receptor-Ns complexes and on receptors uncoupled from Ns.
The effects of Mg2+ or ethylenediaminetetraacetic acid (EDTA) on 125I-glucagon binding to rat liver plasma membranes have been characterized. In the absence of guanosine 5'-triphosphate (GTP), maximal binding of 125I-glucagon occurs in the absence of added Mg2+. Addition of EDTA or Mg2+ diminishes binding in a dose-dependent manner. In the presence of GTP, maximal binding occurs in the presence of 2.5 mM Mg2+ (EC50 = 0.3 mM) while EDTA or higher concentrations of Mg2+ diminish binding. Response to exogenous Mg2+ or EDTA depends on the concentration of Mg2+ in the membranes and may vary with the method used for membrane isolation. Solubilized 125I-glucagon-receptor complexes fractionate on gel filtration columns as high molecular weight, GTP-sensitive complexes in which receptors are coupled to regulatory proteins and lower molecular weight, GTP-insensitive complexes in which receptors are not coupled to other components of the adenylyl cyclase system. In the absence of GTP, 40 mM Mg2+ or 5 mM EDTA diminishes receptor affinity for hormone (from KD = 1.2 +/- 0.1 nM to KD = 2.6 +/- 0.3 nM) and the fraction of 125I-glucagon in high molecular weight receptor-Ns complexes without affecting site number (Bmax = 1.8 +/- 0.1 pmol/mg of protein). Thus, while GTP promotes disaggregation of receptor-Ns complexes, Mg2+ or EDTA diminishes the affinity with which these species bind hormone. In the presence of GTP, hormone binds to lower affinity (KD = 9.0 +/- 3.0 nM), low molecular weight receptors uncoupled from Ns.(ABSTRACT TRUNCATED AT 250 WORDS)
Lipson
K E
KE
Memorial Sloan-Kettering Cancer Center, New York, New York 10021.
Kolhatkar
A A
AA
Maki
R G
RG
Donner
D B
DB
D000818:Animals; D002413:Cations, Divalent; D002462:Cell Membrane / Q000378:metabolism; D004492:Edetic Acid / Q000494:pharmacology*; D005934:Glucagon / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008274:Magnesium / Q000494:pharmacology*; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D011964:Receptors, Gastrointestinal Hormone / Q000187:drug effects / Q000378:metabolism*; D018027:Receptors, Glucagon
D002413:Cations, Divalent; D011964:Receptors, Gastrointestinal Hormone; D018027:Receptors, Glucagon; D006160:Guanosine Triphosphate; D005934:Glucagon; D004492:Edetic Acid; D008274:Magnesium