I have a question about describing the protein motion amplitude in 3D Flex. When I performed 3D flex generation, I noticed that in one state, such as substrate binding to the protein, the motion amplitude or dynamics in the catalytic region of the enzyme became smaller than in the state without substrate. However, this is only observed by eyes. I am wondering if there is some way that I can describe the motion difference through quantification.
In the current state of the technology, I recommend using the latent variables to make same-size groups of particles along the motion trajectory in question, separately reconstructing them, fitting models to each, and then making measurements straightforwardly on the ensemble of atomic models.
@DanielAsarnow makes a great suggestion! Those refined volumes could be modeled as multiple states using a tool such as varref in Phenix. From there you might be able to infer information about the magnitude of motions that you are interested in understanding.
If comparing between different datasets (e.g. substrate-bound vs substrate-free) it is worth considering that particle curation, as well as image and sample quality can differ from dataset to dataset (or processing carried out person to person) and may affect the results of 3DFlex or 3DVA. It’s highly recommended that differences in the magnitude of observed motions be validated by orthogonal biochemical or biophysical approaches such as HDX-MS, NMR (NOE or HDX), and nanoDSF, or by mutagenesis studies once you have developed models/hypotheses.
Thank you DanielAsarnow, my map is not a high resolution map, which makes me worry that it may not be fully trusted when I use it to build a model. But I can try to improve the map and holp it can reach to 3~4 A resulution.
Thanks so much!
Hi kstachowski, thank you for the detailed suggestion. I will try other approch to figure out the dynamic motion of the protein in different state. Thanks so mcuh!