Re: [ccp4bb]: Riding Hydrogens

[Bart Hazes <bhazes@ualberta.ca>]

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On Fri, 6 Sep 2002, Ethan Merritt wrote:

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> On Friday 06 September 2002 11:30, you wrote:
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> > Surprisingly, I found that adding the hydrogens in lowered both R and
> > R free by about 1%.
> > So I am unclear on on the requirements for adding the hydrogens on in
> > refinement. At what resolutions do they become justified? Why are
> > they lowering my R and R-free?
>
> This question is a bit more subtle than you may think.
> Since hydrogen is a weak scatterer, it only contributes significantly at
> low resolution (yes - low not high).  So if you can place all those H
> electrons at their correct position in the cell then your R and Rfree
> should improve even at quite low resolution.  The difficulty is in that
> "correct position" bit.  Usually it is only when your structure is well
> refined to high resolution (better than 2A let's say) that the heavy atom
> positions are accurate enough that you have a good shot at placing
> the hydrogens correctly.
>
> So if you take a model refined at atomic resolution and calculate
> R factors against 3A data with and without hydrogens, it is likely
> you will see an improvement from including the hydrogen scattering.
> But if you only had 3A data to begin with for that same structure,
> it is unlikely that you would have been able to generate a model
> accurate enough to benefit from adding hydrogens.  Not because
> the hydrogen scattering is insignificant, but because you cannot
> place the hydrogens correctly.

Since they are riding hydrogens, I would expect that the coordinate error in
the hydrogens is similar to that of the carbon and nitrogen atoms to which
they are attached. Clearly, the coordinate error will be greater for a lower
resolution structure but at the same time, coordinate errors have less impact
on R/Rfree at lower resolution. So I would think adding hydrogens still
improves the model even though the effect on R/Rfree will be less.

People should also keep in mind that adding 8 hydrogens and therefore 8
electrons does not have the same effect as adding 1 oxygen that also has 8
electrons. The effect is only the same if all 8 hydrogen electrons scatter in
phase and of course they don't. You actually have to add 64 (8**2) hydrogens
to get the effect of 1 oxygen (probably even more since, as Ethan pointed out,
the atomic scattering factor for hydrogen falls off faster with resolution).

Another practical point concerns the treatment of the hydrogen atoms by the
refinement program. Are they used for pairwise interaction calculations, are
they used to determine the solvent mask. If so that can effect the refinement
as well.

>klaas@ultr.vub.ac.be wrote
> But what does it mean. In terms of enzymatic activity, substrate
> specificity, entropy-enthalpie balance, surface properties, crystal
> properties, beam line setup, geological stability of the storage ring, ...
> (And how to convince the referees).

It just means that you place hydrogens where you already knew they would be.
You don't gain new knowledge, you just visualize them and, by reducing
R/Rfree, you should get a slighly cleaner electron density map. No need to
convince referees of anything. Unfortunately, hydrogens involved in catalysis
tend not to be typical riding hydrogens.

Bart

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