Youre studying cellular signaling through G-Protein Coupled Receptors (GPCRs). Specifically youre working on a pair of newly identified GPCRs GPCR-A and GPCR-B. Each binds the same small ligand but activates different heterotrimeric G-proteins that act on adenylyl cyclase.
A (3 points). Your grad student mentor explains that both the receptors and the G and G subunits of the heterotrimeric G-proteins are membrane associated. Youre puzzled because although you find obvious transmembrane domains in the receptor you find none in G or G. How are G and G associated with the membrane (be specific)?
B (5 points). You examine the sequences of the seven predicted transmembrane -helices of GPCR-B. Youre surprised to find that there are hydrophilic amino acid residues in these helices. How can you explain this observation?
You find that the binding of ligand has opposite effects on adenylyl cyclase activity for each receptor. GPCR-A causes an increase in adenylyl cyclase activity while GPCR-B causes a decrease in adenylyl cyclase activity.
C (2 points). Based on the above stated effects on adenylyl cyclase activity what type of G-protein is activated by each receptor?
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Name: ______________________________D (6 points). You have a cell line that expresses both GPCR-A GPCR-B the
corresponding G-proteins and adenylyl cyclase. There is a basal level of adenylyl cyclase activity that produces a baseline cAMP concentration. Your project is to characterize a series of mutations in these components. Will each mutation increase or decrease the intracellular levels of cAMP upon ligand addition (remember both GPCR-A and GPCR-B bind the same ligand)?
A mutation in GPCR-A that prevents G- protein activation?
A mutation in GPCR-B that prevents G- protein activation?
A mutation in Gs that prevents release of bound GDP.
A mutation in Gi that prevents release of bound GDP.
A mutation in Gs that prevents GTP hydrolysis.
A mutation in Gi that prevents GTP hydrolysis.