The influence of monovalent cations on trimeric G protein Gi1α activity in HEK293 cells stably expressing DOR-Gi1α (Cys-Ile) fusion protein

Short title: Inhibition of δ-opioid receptor (DOR) signaling by monovalent ions 2 Summary The effect of monovalent cations on trimeric G protein G i 1α was measured at equimolar concentration of chloride anion in pertussis-toxin (PTX)-treated HEK293 cells stably expressing PTX-insensitive DOR-G i 1α (Cys 351-Ile 351) fusion protein by high-affinity [ 35 S]GTPγS binding assay. The high basal level of binding was detected in absence of DOR agonist and monovalent ions and this high level was inhibited with the order of: Na + > K + > Li +. The first significant inhibition was detected at 1 mM NaCl. The inhibition by monovalent ions was reversed by increasing concentrations of DOR agonist DADLE. The maximum DADLE response was also highest for sodium and decreased in the order of: Na + > K + ≈ Li +. Our data indicate i) an inherently high activity of trimeric G protein G i 1α when expressed within DOR-G i 1α fusion protein and determined in the absence of monovalent cations, ii) preferential sensitivity of DOR-G i 1α to sodium as far as maximum of agonist response is involved.