Following their activation upon binding to ligand, G protein coupled receptors,
ID: 164559 • Letter: F
Question
Following their activation upon binding to ligand, G protein coupled receptors, GPCRs, in turn activate G proteins and thereby initiate a signaling cascade. Hollowing ligand binding. GPCRs become phosphorylated by G protein receptor kinases (GRKs) which, in turn, leads to desensitization of the receptor such that continued stimulation by ligand results in a waning responsiveness of the cell. Arrestins are proteins that bind to GPCRs and are involved in this desensitization. In order to understand GPCR-arrestin interactions, the beta_2-adrenergic receptor (beta_2-AR), a GPCR, and its interaction with beta arrestin are subjected to study. A cell line expressing the beta_2-AR is incubated in epinephrine, a ligand for this receptor, for S minutes. The cells arc then lysed and the beta_2 - AR is immunoprecipitated from the lysate. To determine if beta arrestin is bound to the receptor. the immunoprecipitate is examined by Western blotting using an antibody directed against beta arrestin. The experiment is repeated, but this time prior to and during epinephrine addition the cells are incubated in an inhibitor that blocks beta_2-AR phosphorylation by the kinase GRK2 (BARK). The data are shown below. a) How does the activation state of the receptor affect beta arrestin binding? b) What information do the kinase inhibitor studies provide about beta arrestin binding to the receptor?Explanation / Answer
a Ans:
b Ans:
Kinase inhibitor studies provide the kinase GRK2 blocks the Beta2-adrenergic receptor phophorylation.
if beta-arrestin is bound to the receptor, the immunoprecipitate is examined by western blotting using an antibody directed against beta-arrestin.