Hi Mischa,
We have a similar case. There is difference density, but only for some of
> the hydrogens (mostly methyl groups on Leu, Ile, Val, Ala). How does one
> decide which hydrogens to include in explicit refinement? The case in
> question has 0.99Å data (diffraction is significantly better, but data were
> collected to only 0.99Å).
>
> Sorry, first time refining at that kind of resolution.
>
this might be helpful:
- some overview of handling H atoms in refinement:
Acta Cryst. (2010). D66, 1153-1163
Joint X-ray and neutron refinement with phenix.refine
- specifics and tricks of refinement at very high resolution:
Acta Cryst. (2007). D63, 1194-1197
On macromolecular refinement at subatomic resolution with interatomic
scatterers
Acta Cryst. (2004). D60, 260-274
On the possibility of the observation of valence electron density for
individual bonds in proteins in conventional difference maps
Acta Cryst. (2010). C66, o585-o588
N-{N-[2-(3,5-Difluorophenyl)acetyl]-(S)-alanyl}-(S)-phenylglycine tert-butyl
ester (DAPT): an inhibitor of [gamma]-secretase, revealing fine electronic
and hydrogen-bonding features
Let me know if you have questions.
Most of the time we don't see H atoms in X-ray maps at typical
"macromolecular" resolutions, but this doesn't mean that H atoms are
disordered or not present in actual structures. It does mean that the model
and/or data quality are not high enough to see them. In a hypothetical case
of perfect model and data you would see hydrogen atoms even in a 3A
resolution map. In fact, hydrogen atoms are weak scatterers so they
contribute the most to low resolution reflections while their contribution
to high resolution data approaches zero (just plot the bin average <F> for H
and non-H as a function of resolution). Say at 2A resolution the maps are
much more noisier than the maps at 0.7A resolution (when we routinely see H
atoms). So getting higher resolution data means getting more accurate model,
which in turn means getting less noise and therefore more features that you
can see.
In summary - if you want to see more hydrogens (or the one you are chasing)
then you need:
1) get higher resolution data (as high as you can),
2) do the best possible job refining your model. Also note, even for Aldose
reductase model refined at 0.66A resolution one could see only ~50-60% of
hydrogens,
3) OR simply collect neutron data (if possible of course).
Cheers,
Pavel.
