Human substance P receptor expressed in Sf9 cells couples with multiple endogenous G proteins

摘要

To identify the G proteins involved in the function of human substance P receptor (hSPR), the receptor was expressed in Sf9 cells using the baculovirus expression system. Maximal hSPR expression was up to 65 pmol/mg membrane protein. The following data indicated that hSPR in Sf9 membranes is coupled to endogenous G proteins: 1) binding of agonist radioligand [I-125]BHSP to the receptor was sensitive to guanine nucleotides; and 2) stimulation of the receptor increased [S-35]GTP gamma S binding. The hSPR-associated G proteins were identified by photoaffinity labeling with [alpha-P-32]-azidoanilido GTP ([alpha-P-32]AAGTP), followed by immunoprecipitation of the labeled G proteins with antibodies specific for various G alpha-subunits. These experiments showed that stimulation of hSPR in Sf9 membranes activated multiple endogenous G proteins including G alpha(o), G alpha(q/11), and G alpha(s). While hSPR's ability to associate with G(q/11) is well-documented, the present study provides the first evidence of hSPR's potential to activate G alpha(o) and G alpha(s). [References: 47]