PS it’s probably not a salt crystal…TCEP is not a salt, your ligand I presume is also not a salt, a small cleaved peptide neither. As to why previously in a very similar condition you did get your desired protein plus (other) ligand crystal, it just means the molecule (TCEP') crystallises in a similar condition to your protein - I don’t think you can conclude much more than that (unless there is some other difference like the TCEP being older this time and more oxidised, for example).
Mark J van Raaij Dpto de Estructura de Macromoleculas, lab 20B Centro Nacional de Biotecnologia - CSIC calle Darwin 3 E-28049 Madrid, Spain tel. +34 91 585 4616 (internal 432092) Section Editor Acta Crystallographica F https://journals.iucr.org/f/ > On 4 Feb 2023, at 15:48, kavyashreem <kavyashr...@instem.res.in> wrote: > > Dear all, > > Sorry for the confusion created, I did not mean that a protein would have fit > in the small unit cell. My question was - > > 1. Why are there closely spaced spots arising in salt crystal? > > 2. If TCEP could crystallize in the condition, I have got a protein (same as > this)+ligand (different ligand) complex in very close condition. (ligand size > is within 500Da). > > Thank you > > Kavya > > > > On 2023-02-03 14:35, a.perra...@nki.nl wrote: > >> Hi Kavya, >> >> Try https://csb.wfu.edu/tools/vmcalc/vm.html >> <https://csb.wfu.edu/tools/vmcalc/vm.html> >> >> This tells you that a 30kD protein simply does not fit the cell. >> >> I am pretty sure you crystallised the ligand, or TCEP actually. >> >> Also, if you look at the diffractions pattern, its clear the crystal >> diffracts beyond 1.0A, diffraction spots are really very very very strong at >> 2.0A. >> >> >> >>> On 3 Feb 2023, at 09:22, kavyashreem <kavyashr...@instem.res.in >>> <mailto:kavyashr...@instem.res.in>> wrote: >>> Dear all, >>> >>> We crystallized a protein (30kDa) + ligand (by cocrystallization), in the >>> condition 10%PEG3350, 50mM Zinc acetate. >>> Protein was in the buffer 20mM HEPES, 150mM NaCl, 1uM ZnCl2, 4mM TCEP, pH >>> 8. >>> Crystal: Crystal: >>> crystal under UV m >>> <b06fc576.png> <e091c7fd.png> <8ef9453e.png> >>> When we collected the data at an in-house facility, it looked something >>> like this: >>> <b903961d.png> >>> The minimum resolution spot is around 9Ang and maximum ~2.2Ang. >>> I have not come across a protein diffraction like this, nor of a salt. When >>> I ran the gel for the incubated protein (protein+ligand), there was no >>> degradation. >>> Although, I was sure there is some problem with this image I tried >>> processing, which could not be, But indexing showed a unit cell of 11Ang, >>> 11Ang, 46Ang in P3. which was quite expected for two of the axes but not >>> the third. >>> Can anyone please shed some light on this diffraction image? >>> How can it happen? >>> >>> Thank you >>> Regards >>> Kavya >>> >>> >>> To unsubscribe from the CCP4BB list, click the following link: >>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 >>> <https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1> >> To unsubscribe from the CCP4BB list, click the following link: >> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 >> <https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1> > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 > <https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1> ######################################################################## 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/
