I have gone through a simple idea that proteins have different conformations; it is a dynamic system. Moreover, whenever we do ensemble experiments, so we get an average picture. There are some questions-:
Q.1 Often protein structures are reported in PDB (protein data bank), so what do they signify? The most stable conformation or some averaged picture of all conformation or both?
Q.2 In the case of the NMR, proteins have the freedom to move/vibrate through space as they are in the solution phase, but in XRD, they are rigid. So what will be the difference if I get one structure from XRD and one from NMR?
Q.3 Why structure of some proteins has not been resolved yet? I get it that a large protein cannot be solved using NMR; it will be much complex. However, why we cannot use XRD? a) Why is the crystallisation of some protein hard? b) Will there be any other challenges even if we get the crystals of a protein?
Q.4 Investigators often use different buffers to crystalise the given protein, but how they make sure that structure that they are getting in that buffer is the "actual" structure? How they make sure that crystalisation do not affect the "actual" structure of the protein.
From "Actual" Structure, I mean the structure or set of conformations that are retained in the physiological condition (in the working conditions).