On Monday, January 07, 2013 12:10:17 pm Edward A. Berry wrote:
> The idea is (whether it's valid or not) to apply the information from
> both sites simultaneously. If the density is pretty ambiguous and one side
> tends to drift off into an alternate conformation and the other drifts off
> into another conformation, but you have every reason to believe that
> the conformation is the same on both sides, applying NCS allows you to
> refine a single conformation that is consistent with both.

Ah, but what if you don't have every reason to believe
that the conformation is the same on both sides?

> More generally, is there any reason to not? I suppose ligands
> may be more likely than amino acids to violate NCS, but good
> practices would say examine each residue for violations.
>
> You could say, why enforce NCS on the 27'th residue of each chain, since
> their contribution to the number of parameters is small.
> (mind you,there may be good reasons to not constrain ligands
> that I m not aware of, if so I hope someone speak up)

The example mentioned earlier was HIV protease (a homodimer) in complex
with an asymmetric inhitor. I.e. a single asymmetric ligand occupying a
single site formed by 2 NCS-related chains.  Depending on what treatment
of NCS is available, it may or may not be reasonable to apply NCS
restraints to the protein.  Local geometry restraints may be
reasonable but coordinate restraints probably not.

In practice you would normally expect the ligand to be stochastically
present in one of 2 orientations, corresponding to the 2-fold NCS.
But if the ligand is not totally enclosed by the protein, the two
orientations may not be equally present and indeed might conceivably
not follow the overall 2-fold NCS of the protein.

To bring in an example that I've worked on myself, the cholera toxin
B-pentamer has 5 identical receptor binding sites related by 5-fold
NCS.  Small-molecule receptor analogs bound at those sites can extend well
beyond the protein envelope; their precise binding conformation is
affected by lattice contacts that obviously don't follow 5-fold NCS.
This is a case where applying the protein NCS to the ligands
would not make sense.


	Ethan



>
> eab
>
> Boaz Shaanan wrote:
> > Just a naive question: why apply NCS to ligands at all? Their contribution to the number of parameters and hence to the param/obs ratio, the main argument for applying NCS, is negligible, isn't it?
> >
> >   Boaz
> >
> > Boaz Shaanan, Ph.D.
> > Dept. of Life Sciences
> > Ben-Gurion University of the Negev
> > Beer-Sheva 84105
> > Israel
> >
> > E-mail: [log in to unmask]
> > Phone: 972-8-647-2220  Skype: boaz.shaanan
> > Fax:   972-8-647-2992 or 972-8-646-1710
> >
> >
> >
> >
> >
> > ________________________________________
> > From: CCP4 bulletin board [[log in to unmask]] on behalf of Edward A. Berry [[log in to unmask]]
> > Sent: Monday, January 07, 2013 6:12 PM
> > To: [log in to unmask]
> > Subject: Re: [ccp4bb] About NCS and inhibitors
> >
> > But I think the original poster meant partially overlapped after applying
> > the ncs- operator- i.e. they are not ncs related but occupy partly the same
> > position (in the two non-overlapping copies of the binding site).
> >
> > Then I guess it depends how clear the density is- If the density is not very
> > clear and if the protein residues of the active site do follow ncs, I would try
> > rebuilding ligand b to match a and (separately) a to match b and refining
> > with ncs applied to the ligand; and see if the resulting fit looks just as good.
> > eab
> >
> > Joel Sussman wrote:
> >> Dear All,
> >> Something like what Felix wrote is seen in the crystal structure of *recombinant human acetylcholinesterase* (*rhAChE*)
> >> (PDB-ID: *3lii*), with two molecules are seen in the asymmetric unit.
> >> * In one molecule, the active-site gorge (where inhibitors normally lie) is occupied with part of a peptide loop from a
> >> symmetrically related rhAChE.
> >> * While the corresponding region of the other copy of rhAChE is void of this peptide.
> >> See figs 15-16 in:
> >> Dvir, H., Silman, I., Harel, M., Rosenberry, T. L.&  Sussman, J. L. (2010). "Acetylcholinesterase: From 3D structure to
> >> function" /Chemico-Biological Interactions/ *187*, 10-22.
> >> * So, in essence, no reason to ever assume that two copies in asymmetric unit will be identical, or have identical
> >> inhibitors bound, or 'surrogate inhibitors' (like in this case) bound. Sometimes differences are due to difference in
> >> crystal packing
> >> Best regards,
> >> Joel
> >>
> >>
> >> On 7 Jan 2013, at 11:58, Felix Frolow wrote:
> >>
> >>> I apologise for typing blinbly:
> >>> " if one is in, the second can't be"
> >>> FF
> >>> Dr Felix Frolow
> >>> Professor of Structural Biology and Biotechnology, Department of Molecular Microbiology and Biotechnology
> >>> Tel Aviv University 69978, Israel
> >>>
> >>> Acta Crystallographica F, co-editor
> >>>
> >>> e-mail: [log in to unmask]<mailto:[log in to unmask]>
> >>> Tel: ++972-3640-8723
> >>> Fax: ++972-3640-9407
> >>> Cellular: 0547 459 608
> >>>
> >>> On Jan 7, 2013, at 11:48 , Felix Frolow<[log in to unmask]<mailto:[log in to unmask]>>  wrote:
> >>>
> >>>> Why not? They can be mutually excluding! If one is in, the second can be. This phenomenon brakes a local symmetry.
> >>>> FF
> >>>>
> >>>> Dr Felix Frolow
> >>>> Professor of Structural Biology and Biotechnology, Department of Molecular Microbiology and Biotechnology
> >>>> Tel Aviv University 69978, Israel
> >>>>
> >>>> Acta Crystallographica F, co-editor
> >>>>
> >>>> e-mail: [log in to unmask]<mailto:[log in to unmask]>
> >>>> Tel: ++972-3640-8723
> >>>> Fax: ++972-3640-9407
> >>>> Cellular: 0547 459 608
> >>>>
> >>>> On Jan 7, 2013, at 11:28 , Xiaopeng Hu<[log in to unmask]<mailto:[log in to unmask]>>  wrote:
> >>>>
> >>>>> Dear All,
> >>>>>
> >>>>> We recently resolved an enzyme/inhibitor complex structure. The enzyme contains two NCS related active site and we
> >>>>> did find extra density in both of them.However we observed that the two inhbitor moleculors are not NCS related, but
> >>>>> partly overlaped if make a NCS moleculor. Has anyone else observed this before? Thanks for any help and suggestion!
> >>>>>
> >>>>>
> >>>>> Best,
> >>>>>
> >>>>> Xiaopeng Hu
> >>>>>
> >>>>
> >>>
> >>
> >
>

--
Ethan A Merritt
Biomolecular Structure Center,  K-428 Health Sciences Bldg
University of Washington, Seattle 98195-7742