Dear Deepak,
I have a few suggestions:
1) First you can send it to synchrotrons, particularly microfocus beamlines
and shoot as many good in-house diffracting crystals.
2) Setting up crystallisation by micro batch method under Al's oil at
various vapour exchange rate may help. But you have to harvest crystals at
the right time.
3) XFEL with micro crystals will also solve your problem.
4) More difficult one is to do MicroED. You need to do a cryoFIB and do
MicroED, ie., diffraction using cryo electron microscopy and do MR with
your available model.
Trial and perseverance will definitely lead you to the structure of your
required complex..
What is the solvent content in your protein crystal? Based on that you can
also plan other trials.
Hope that helps.
Wish you all the very best.
Natesh
On Tue, 18 May 2021 at 15:49, Deepak Deepak <deepmalik...@gmail.com> wrote:
> Dear all,
>
> I have got multiple crystals (see picture 1) of a protein (8kDa) with a
> helical aromatic oligoamide foldamer (5kDa) but these crystals *diffract
> very poorly *(see the diffraction pattern in picture 2).
>
> I prepare a 1.3mM:1.3mM complex of protein: foldamer in 20mM Tris, pH 7.5
> buffer. Crystals grew in 3-5 days in sitting and hanging drop at 20 Deg C
> and 25 Deg C in the following conditions:
>
> *- 20% PEG 400, 0.1M MES pH 6.0*
> *-20% PEG 400, 0.1M Sodium Cacodylate pH 6.0*
>
> *Multiple cryo used were:*
> *-25%Glycerol in mother solution*
> -30% glycerol in water
> *-30%PEG 400,*
> *-35% PEG 400*
> *-20% PEG 8000 + 40% PEG 400 mix*
>
> Kindly suggest some methods/modifications on how can I improve the
> resolution and get better-diffracting crystals. Please let me know if you
> need more information.
>
> Kind regards,
> Deepak
> Ph.D. Student
>
> PS: The protein is a DNA binding protein and I have crystallized and
> solved the structure of this protein with its DNA partner and now I
> crystallized it with our foldamers but diffraction is not good. There are
> multiple structures of the Protein+DNA complex in literature but *no
> apo-protein structure *as the protein needs a binding partner to
> crystallize. *We already have solution studies showing a good binding.*
>
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--
----------------------------------------------------------
"Live Simply and do Serious Things .. "
- Dorothy Mary Crowfoot Hodgkin OM, FRS
"In Science truth always wins"
- Max Ferdinand Perutz OM FRS
----------------------------------------------------------
Dr. Ramanathan Natesh
Assistant Professor,
School of Biology,
Indian Institute of Science Education and Research Thiruvananthapuram
(IISER-TVM),
Maruthamala P.O., Vithura,
Thiruvananthapuram, 695551, Kerala, India
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