Needs to be done by tonight 12:00 A.M today
California
Redwood city
You are using cryo-EM to study a very large membrane receptor. This protein has 5 domains named A-E. Domains A-D are known to be tightly imbedded in the membrane and Domain E is a known cytosolic domain that binds to A-D upon ligand binding. You are studying this protein embedded in Nanodiscs and in the presence of the ligand. In your initial round of 3D reconstruction (after 2D and 3D classification), Domain E resolves to only about 8 Å, whereas all of the other features of the protein resolve to < 4.0 Å. Based on experimental data, you suspect that Domain E can bind in multiple conformations to Domains A-D. What next steps could you do in the data processing to better resolve Domain E? What do you expect the outcome to be if your suspicions prove correct?
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