Hi Oliver,
Hello,
the ordered_solvent option in phenix.refine apparently uses these parameters by default:
min_model_peak_dist = 1.8 max_model_peak_dist = 6.0 min_peak_peak_dist = 1.8
I was wondering about the justification for this (extremely relaxed) set of limits. As far as I know, the shortest hydrogen bonds (if defined as donor-acceptor distance) are about 2 A, and even these are considered exceptional and rare. Similarly, anything in excess of 3.5 A is quite unlikely to be significant energetically, so why use 6 A as an upper limit? Even if max_model_peak_dist was only meant to limit the extent of the outer solvent shell, there is no such thing as max_peak_peak_dist to avoid unreasonably long individual bonds. My personal suggestion would be a range of about 2.3 - 3.2 A for a significant hydrogen bond, with the lower limit exemplified by charged interactions (carboxyl-water or oxonium-water), which tend to be shorter - and stronger - than average.
these are the parameters that you can cast them to match your taste. "max_model_peak_dist=6.0" is the maximum distance that a water is allowed to be from the macromolecule. However, this doesn't mean that the water can freely float at 6A or so distance from the macromolecule, since the other restrictions (h_bond_max = 3.2) will make sure that this water is bridged to the macromolecule or another water. Note, the parameters you referred to min_model_peak_dist = 1.8 max_model_peak_dist = 6.0 min_peak_peak_dist = 1.8 are used to initially select peaks, and the other set of parameters h_bond_min_mac = 1.8 h_bond_min_sol = 1.8 h_bond_max = 3.2 is used to make further selection. The next level of selection is the map correlation between 2mFo-DFc and DFc maps computed for each water (the default threshold is 0.7 - that is if the map CC is lower than 0.7, the peak-candidate is discarded). There are bunch of other selection criteria as well. I agree, 1.8 is a generous choice, but given the amount of (many) other selection criteria that phenix.refine uses to distinguish water from non-water, I hope in the end the noise gets filtered out. On the other hand, we all know that we never work with the exact electron density but with its Fourier image that is always distorted due to finite resolution and data completeness (plus other data and model errors). These kinds of distortions may perturb water peak position in both directions. The shorter distance cutoff (1.8A) is meant to account for this in order to not miss such waters.
On the other hand, I do frequently observe elongated densities which, in accordance with the default parameters, are typically interpreted by phenix.refine as two water molecules about 2 A apart. So the question arises whether this is a realistic assumption, compared to a "sliding water" model in which the position of a single molecule is just ill-defined in a certain direction (due to a competition of adjacent binding sites for example)?
This kind of situations is best modeled using waters in alternative conformations (if I correctly understood what you mean), which is not implemented yet in phenix.refine - phenix.refine does not build waters in alternative conformations automatically). So I guess placing two waters is probably not ideal, but still is better than nothing. Ideally it would be better to assign one water altloc A and the other one altloc B, so their occupancies are constrained to 1. Please let me know if you have a specific example showing that phenix.refine placed wrong waters, and I will be happy to have a closer look. All the best! Pavel