Hi,
I'm attempting to run ResolveCryoEM, and I have a few questions about setting the job up correctly. My half-maps were reconstructed with C4 symmetry to ~2 Å (CryoSPARC), and my model was refined into the density with the "Ncs constraints" box checked (Phenix RSR). My sample is a protein-DNA complex (4 copies of a protein protomer, 4 copies of a ssDNA protomer). I'm running Phenix 1.21.1-5286-000 (the latest installation provided by SBGrid).
In the first version of the run, I only supplied the two half-map files and a seq file.
Question 1:
My current seq.dat file looks like this (as mentioned previously in the forum, I was getting errors when I tried the version where my seq.dat file has only 1 protomer and I set the number of copies to 4 in the GUI field "Copies of sequence file in map"):
>protein_protomer1
MGSRSG...
>protein_protomer2
MGSRSG...
>protein_protomer3
MGSRSG...
>protein_protomer4
MGSRSG...
Currently, Phenix is reading in the "AAAAAA" as a protein chain ("Chain types considered: PROTEIN"). How do I tell Phenix that those are deoxyadenosines, not alanines?
Question 2:
Is Phenix automatically detecting the C4 symmetry and density-modifying with that constraint? The resulting map looks symmetric in the noise, but I don't see anything in the log suggesting that it knows the symmetry. Here are some settings from the log that seem relevant (all left to default values):
refine_symmetry_in_denmod = True
use_symmetry_in_denmod = False
symmetry_in_denmod_last_cycle_only = True
fraction_ncs = None
I attempted to supply a symmetry file (the output of a MapSymmetry job on the full map = "symmetry_from_map.ncs_spec"), but the file fails to load into the input ("Phenix error: Phenix did not recognize the file type for ****/symmetry_from_map.ncs_spec").
Question 3:
A part of my protein is disordered and does not appear in the cryo-EM density (and was thus left unmodeled). Is it correct to leave that piece out of the seq.dat file?
Question 4:
If I include a model in the input, Phenix demands a symmetry file, but I couldn't get my symmetry file to load as explained above. What is the symmetry file supposed to look like?
Thank you!
-Josh