Unambiguous restraints violation

HADDOCK
1

Hello, I obtained distance restraint information from CX-MS, so I used these unambiguous restraints on HADDOCK. However, the results turned out that the distance between the specific residues violated one of the restraints. Could you please tell me why and how to solve the problem? Thanks.

2

Is it heavily violated?

There can be several issues:

  1. It simply can not be satisfied (e.g. due to required conformational changes)

  2. It is a false positive cross-link

  3. the restraint was wrongly defined and not used (we would need to have the web link of your run to check that - or check for example the it0 out file from the full tar archive, looking for NOE statements and checking if atoms were selected for your specific restraint

3

It is heavily violated. For example, the distance restraint I set is 30 A, but the resulted distance is about 80 A.
The web link of my run is HADDOCK results page (uu.nl). Could you please help me check that? Thank you so much.

4

The unambiguous restraints you defined have an upper limit of 25A, but are defined within molecule A
I.e. they do not influence the docking …

Typo? Shouldn’t you have a segid B in one of the atom selection?

ASSFIL: file unambig.tbl opened.

assign (segid A and resid 323) (segid A and resid 65) 25 25 0
SELRPN: 13 atoms have been selected out of 6730
SELRPN: 8 atoms have been selected out of 6730
assign (segid A and resid 323) (segid A and resid 87) 25 25 0
SELRPN: 13 atoms have been selected out of 6730
SELRPN: 9 atoms have been selected out of 6730
assign (segid A and resid 466) (segid A and resid 76) 25 25 0
SELRPN: 13 atoms have been selected out of 6730
SELRPN: 9 atoms have been selected out of 6730

5

I referred to this paper “Structural prediction of protein models using distance restraints derived from cross-linking mass spectrometry data”.

https://doi.org/10.1038/nprot.2017.146

assign (resid and segid <1st molecule chain>) (resid and segid <2nd molecule chain>)

The first molecule has only one chain, namely chain A. The second molecule also has only one chain, that is chain A. I don’t know which way is correct.

Thank you for answering me.

6

For HADDOCK, each molecule is assigned a different chain and your restraints must reflect that.