(25 POINTS) QUESTION 1. The M3 muscarinic receptor (M3R) signals by activating G
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(25 POINTS) QUESTION 1. The M3 muscarinic receptor (M3R) signals by activating Gq heterotrimers. To determine if M3R associates with Gq before the receptor is activated the M3R was labeled at the N terminus with enhanced cyan fluorescent protein (ECFP) (called C-M3R), and then measured the lateral mobility of Ga labeled with yellow fluorescent protein Venus (Gaa-V). To test membrane mobility HEK293 cells were transfected with C-M3R and GaV, and both proteins were localized to the plasma membrane (Fig 1A Prebleach). A third protein, M4 muscarinic receptor, which links to Ga and Gao, instead of Gag To make a negative control you cross-link the C-M3R and C-TM in the membrane. When examining the data below, what conclusion do you make regarding the ability of the C-M3R to recover after photobleaching? 5 s 100 s C-M3R b 1.0 0.8 0.6 O 0.4 C 1.0 0.8 0.6 0.4 0.2 C-TM C-M3R C-M4R C-TM -C-M3R C-M4R 0.2 60 120 0 60 120 180 Time (s) Time (s)Explanation / Answer
Photobleaching refers to loss of color when illuminated by light.
When C-M3R is labelled with cyan fluorescent protein , then the photobleaching is very high as it can be seen in the image above , that after prebleach the color started disappearing.The same observation can be made by seeing the graph as there is a large dip.
When its lateral mobility was measured with Gaq, that was labelled with yellow fluorescent protein , the effect of photobleaching was not so high as compared with cyan fluorescent protein. As when seeing the graph for venus intensity , the dip was not so deep as when compared with ECFP intensity.
So, concluding this- C-3MR was more photobleached when labelled with yellow fluoroscent protein a compared to when labelled with cyan fluorescent protein.
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