Hi Deepak In my opinion, rMMS microseeding, ie adding crushed seed crystals into *random screens*, should be used routinely in most crystallization projects. It gives much more control, in particular you can control the number of crystals per drop.
In a case like this, I would use it straight away - use the crystals that you showed us to make a seed-stock. The protein is a DNA binding protein and I have crystallized and solved > the structure of this protein with its DNA partner All seed-stocks are not the same - one may work but another may not. Or one particular seed-stock may give crystals with a different unit cell or space group. Since you have a *family *of constructs, you should try cross-seeding with all the other (related) crystals that you have. At the beginning of the project, to keep the number of experiments down, you can mix together several different seed-stocks. However it is recommended that you keep crystals where the main precipitant was PEG separate from crystals where the main precipitant was salt. https://doi.org/10.1107/S2053230X14012552 https://doi.org/10.1107/S0907444907007652 or google MMS or rMMS. Good luck, Patrick On Tue, May 18, 2021 at 11:19 AM 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.* > > ------------------------------ > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 > -- patr...@douglas.co.uk Douglas Instruments Ltd. Douglas House, East Garston, Hungerford, Berkshire, RG17 7HD, UK Directors: Patrick Shaw Stewart, Peter Baldock, Stefan Kolek http://www.douglas.co.uk Tel: 44 (0) 148-864-9090 US toll-free 1-877-225-2034 Regd. England 2177994, VAT Reg. GB 480 7371 36 ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list hosted by www.jiscmail.ac.uk, terms & conditions are available at https://www.jiscmail.ac.uk/policyandsecurity/
