Dear Misba,
A wild guess: have you considered the possibility that this extra
density could be a cacodylate adduct? Cacodylate is well known to react with
thiols - see
https://febs.onlinelibrary.wiley.com/doi/pdfdirect/10.1016/0014-5793(72)80224-2
Here the chemistry is different but you never know. If your data are
redundant enough that you have good anomalous completeness, and were
collected above the As K-edge (11.8667 keV), it might be a good idea to
compute an anomalous difference Fourier and check for the presence of a peak
at the same location as the highest one in your ordinary difference map.
Only a wild guess, though ... .
With best wishes,
Gerard.
--
On Sat, Jan 29, 2022 at 12:18:58PM +0100, Misba Ahmad wrote:
> Placing a water molecule satisfies most of the density and forms nice
> H-bonds but there is still some residual density left (8.6 and 5.6 rmsd).
>
> Best
> Misbha
> [image: 3.png]
>
>
> On Sat, Jan 29, 2022 at 11:05 AM Klaus Futterer <k.futte...@bham.ac.uk>
> wrote:
>
> > Looks more like water molecules. Phosphorylation would give a much bigger
> > peak, and shape of density does not fit either. I don't think this is a
> > covalent modification. Model some water molecules and see what the
> > distances are and what difference density is left.
> >
> >
> > Klaus
> >
> >
> > =======================================================
> > Klaus Fütterer, PhD
> > Reader in Structural Biology
> >
> >
> > School of Biosciences
> > LES College Email:
> > k.futte...@bham.ac.uk
> > University of Birmingham Phone: +44 - 121 - 414
> > 5895
> > Birmingham, B15 2TT, UK (voice mail messages
> > will forward to my email inbox)
> >
> > My normal working hours are Mon - Fri 8.30 - 5.30 pm.
> >
> >
> > =======================================================
> > ------------------------------
> > *From:* CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of
> > misba.ah...@gmail.com <misba.ah...@gmail.com>
> > *Sent:* 29 January 2022 09:45:24
> > *To:* CCP4BB@JISCMAIL.AC.UK
> > *Subject:* Re: [ccp4bb] Help with interpreting Tyrosine modification
> >
> > Hi Tom,
> > The protein was expressed in E Coli.
> >
> > Best
> > Misbha
> >
> > On Sat, 29 Jan 2022, 10:18 Peat, Tom (Manufacturing, Clayton) <
> > tom.p...@csiro.au> wrote:
> >
> >> Hello Misba,
> >>
> >> Doesn't quite look like a phosphate, maybe O-sulfation?
> >> Maybe just as important as the buffer and crystallisation conditions
> >> would be how it was expressed? Insect cells?
> >>
> >> Best of luck, tom
> >>
> >> Tom Peat, PhD
> >>
> >> Biomedical Program, CSIRO
> >> tom.p...@csiro.au
> >>
> >> ------------------------------
> >> *From:* CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of Misba
> >> Ahmad <misba.ah...@gmail.com>
> >> *Sent:* Saturday, January 29, 2022 8:12 PM
> >> *To:* CCP4BB@JISCMAIL.AC.UK <CCP4BB@JISCMAIL.AC.UK>
> >> *Subject:* [ccp4bb] Help with interpreting Tyrosine modification
> >>
> >> Hi all,
> >> I am trying to interpret this strong difference density peak (11.33 rmsd)
> >> that shows up on the tyrosine residue. Any help would be greatly
> >> appreciated.
> >>
> >> Purification buffer: 20mM HEPES pH 7.5, 250mM NaCl, 1mM TCEP, 5mM DTT
> >> Crystallisation condition: Sodium propionate, Sodium cacodylate, BIS-TRIS
> >> propane, PEG 1500
> >>
> >> Best
> >> Misbha
> >> [image: Picture1.png]
> >> [image: Picture2.png]
> >>
> >> ------------------------------
> >>
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