[phenixbb] Low resolution refinement

Ashu Sharma ashuthesharma13 at gmail.com
Tue Nov 6 09:56:36 PST 2018

Thanks so much Pavel !

I already went through placing the crystal structures manually in the
density map and building the entire model. However, I didn't use the
reference model restrains during the refinement and that explains why every
time I refine, the geometry goes bad. I'll do this as you suggested.


On Tue, Nov 6, 2018 at 3:47 AM Pavel Afonine <pafonine at lbl.gov> wrote:

> Hi Ashu,
> you can use phenix.dock_in_map to dock known pieces (from crystal
> structures) into cryo-EM map.
> Then you can use phenix.real_space_refine to refine placed model into
> cryo-EM map. It is important that you use reference model restraints
> with high-res X-ray model as a reference.
> What to do with side chains is a good question. I doubt you can see any
> side chains in 7A resolution map. Reference X-ray model is perhaps your
> best source of knowledge about the side chains. Though keeping them in
> the model in the absence of support from the data seems questionable. I
> doubt there is a clear cut consensus/answer to this.
> You can vary the strength of reference model restraints do vary the dose
> of bias from the known (reference) model.
> Pavel
> > Dear All,
> >
> > I am refining a ~7A single particle cryo-em structure of a protein.
> > The protein has two domains and high-resolution crystal structures of
> > individual domains are already known. I could fit these two crystal
> > structures in the low resolution cryoem map of the full-length
> > protein. I have two questions:
> >
> > 1) Is it acceptable to refine with side chains in this case? In most
> > of the cases, with the guidane from crystal structures, side chains
> > could be placed with reasonable confidence, particularly smaller ones.
> > Should I restrict to poly-ala model or take the advantage of crystal
> > structures and include side chains also?
> >
> > 2) When I tried to refine with side chains, longer side chains like K,
> > R, Q, E which have weaker density tend to bend towards the backbone
> > density. Is there a way to prevent this happening?
> >
> > Suggestions/comments from the community would be highly appreciated.
> >
> > Thanks !
> >
> > Ashu
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