C4 zinc finger protein

[sarah]

Hi, I have been trying to make a haddock model of a zinc finger protein and I need to restrain the cysteine residues to the zinc atom. I have created a tbl file with distance restraints but one of the 4 cysteines does not appear to dock correctly and I’m unsure how to fix this. Also, in my models the cysteine-zinc models appear to be covalently linked (ie in pymol the zinc ion is attached to the cysteine), am I doing something wrong?
Sarah

[amjjbonvin]

Hi Sarah

Remember that the HADDOCK forum is searchable. You will need to restrain your zinc coordination. See

Dealing with elemental ions

The way to define ions for HADDOCK is explained in Box 3 of our Nature Protocols 2010 server paper: The HADDOCK web server for data-driven biomolecular docking. Nature Protocols, 5, 883-897 (2010). Download the final author version here. In short: the residue name should contain the absolute charge state the atom name should have the exact charge E.g. for a zinc atom: residue name: ZN2 atom name: ZN+2 (atom name with four characters should be shifted by one column to the left)

And the way to define distance restraint to the Zinc is explained in the following thread:

Zinc finger protein

Which kind of restraints did you define to maintain the geometry of the zinc finger? To get proper results make sure to define distance restraints between: all atoms coordinating the zinc (basically define the tetrahedron connecting them each coordinating atom and the zinc atom. Note that the distance restraints must have the zinc atom name between double quotes because of the + in the name. The correct nomenclature for HADDOCK for the zinc is: residue name: ZN2 atom name: ZN+2 (shifted …

[sarah]

Hi Alexandre, thanks for your reply. I’m aware that the forum is searchable and have already used both of these references when creating my restraints file. I mapped the distances using NMR data as follows:

assign (resid 295 and segid A and name SG) (resid 298 and segid A and name SG) 3.8 3.8 3.8
assign (resid 298 and segid A and name SG) (resid 316 and segid A and name SG) 3.8 3.8 3.8
assign (resid 295 and segid A and name SG) (resid 316 and segid A and name SG) 3.7 3.7 3.7
assign (resid 295 and segid A and name SG) (resid 319 and segid A and name SG) 3.8 3.8 3.8
assign (resid 316 and segid A and name SG) (resid 319 and segid A and name SG) 3.7 3.7 3.7
assign (resid 298 and segid A and name SG) (resid 319 and segid A and name SG) 3.5 3.5 3.5
assign (resid 295 and segid A and name SG) (resid 1 and segid Z and name “ZN+2”) 2.7 2.7 2.7
assign (resid 298 and segid A and name SG) (resid 1 and segid Z and name “ZN+2”) 2.7 2.7 2.7
assign (resid 316 and segid A and name SG) (resid 1 and segid Z and name “ZN+2”) 2.7 2.7 2.7
assign (resid 319 and segid A and name SG) (resid 1 and segid Z and name “ZN+2”) 2.7 2.7 2.7

As I said, 3 of the 4 cysteines do maintain their position in the tetrahedron, but one doesn’t which is why I’m not sure what I’ve done wrong. Would I need to create additional restraints?

Thanks
Sarah

[amjjbonvin]

Hi Sarah

Your distance restraints are very loose! Probably you misunderstood the meaning of the three numbers:

• first is target distance
• second is the lower limit correction
• third is the upper limit correction

The distance range is this:

Lower limit: first-second
Upper limit: first+third

So in your restraints, e.g. the first one, the distance range is between 0 and 7.6A

[sjenatt]

Hi Sarah,
I am trying to dock zinc to my C2H2 finger, but I can’t get the zinc to the active sites. It’s not even closer to the active site. I am checking if you’ve any suggestions on how to fix this.

When I use active residue, the zinc ion I am using seems to be placed near the active site suggesting there is no problem with the zinc topology I am using for HADDOCK docking.

But, when I am using unambiguous restraints, the zinc ion is >10 nm away from the active site.

I would appreciate any suggestions or comments.

Best,
Sj

[amjjbonvin]

This probably means there is an issue with your restraints file. Please share the iD of a problematic run

[sjenatt]

Hi,
Please see the HADDOCK ID below.

HADDOCK ID:
2701304514

Thanks!

[sjenatt]

I am checking if you get a chance to look at the restraint file. Are there any issues with the restraint file?

[amjjbonvin]

The problem with your run comes from the non ASCII double quotes you used around the zinc atom name in the restraint file.
Because of that the restraints are not properly processed.

[sjenatt]

Thank you so much for finding sim time to look at the issue. I truly appreciate it.

I will rerun using the corrected ascii format.

Best regards,
Sj