Phosphate and thiophosphate group donating adenine and guanine nucleotides inhibit glibenclamide binding to membranes from pancreatic islets

Summary

In microsomes obtained from mouse pancreatic islets, the Mg complex of adenosine 5′-triphosphate (MgATP) increased the dissociation constant (K _D) for binding of [³H]glibenclamide by sixfold. In the presence of Mg²⁺, not only ATP but also adenosine 5′-0-(3-thiotriphosphate) (ATPγS), adenosine 5′-diphosphate (ADP), guanosine 5′-triphosphate (GTP), guanosine 5′-diphosphate (GDP), guanosine 5′-0-(3-thiotriphosphate) (GTPγTS) and guanosine 5′-0-(2-thiodiphosphate) (GDPβ S) inhibited binding of [³H]glibenclamide. These effects were not observed in the absence of Mg²⁺. Half maximally effective concentrations of the Mg complexes of ATP, ADP, ATPγS and GDP were 11.6, 19.0, 62.3 and 90.1 μmol/l, respectively. The non-hydrolyzable analogues adenosine 5′-(β,γ-imidotriphosphate) (AMP-PNP) and guanosine 5′-(β,γ-imidotriphosphate) (GMP-PNP) did not alter [³H]glibenclamide binding in the presence of Mg²⁺. MgADP acted much more slowly than MgATP and both MgADP and MgADP did not inhibit [³H]glibenclamide binding when the concentrations of MgATP and MgATP were kept low by the hexokinase reaction. Development of MgATP-induced inhibition of [³H]glibenclamide binding and dissociation of [³H]-glibenclamide binding occurred at similar rates. However, the reversal of MgATP-induced inhibition of [³H]glibenclamide binding was slower than the association of [³H]glibenclamide with its binding site. Exogenous alkaline phosphatase accelerated the reversal of MgATP-induced inhibition of [³H]glibenclamide binding. MgATP enhanced displacement of [³H]glibenclamide binding by diazoxide. The data suggest that sulfonylureas and diazoxide exert their effects by interaction with the same binding site at the sulfonylurea receptor and that protein phosphorylation modulates the affinity of the receptor.