[ccp4bb] Unknown electron density
Dear CCP4BB, I am working on a structure refinement and came across a blob of density that looks like an anvil (see link). Has anyone seen this before and have an idea of what it is? https://drive.google.com/file/d/1BGNwV3MujYjrV4W5m1bh3_WWd49RhJ1w/view?usp=sharing The protein is in potassium phosphate and the components in the crystallization/cryo condition are zinc acetate, PEG, and glycerol. Thank you in advance for your time and help! Betty To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=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/
Re: [ccp4bb] AW: [ccp4bb] Unknown positive electron density
Thank you all for your replies and suggestions. I have fitted in some dummy waters as Herman suggested and this is the result after one round of refinement. The waters are positioned where there is highest electron density, but I am not sure how to move forward from here. https://cbsostorage.chem.umd.edu/owncloud/index.php/s/VTQ6Uinp1UVIWGR https://cbsostorage.chem.umd.edu/owncloud/index.php/s/NUZKClayzSMn7PP https://cbsostorage.chem.umd.edu/owncloud/index.php/s/0FZhxtgcQhhqnaA I have also tried fitting in a nucleotide, but the geometry of it does not fit the density well. We do not think this is due to Fourier truncation ripples since the positive density is not at the crystallographic symmetry axis. Betty On Tue, Aug 22, 2017 at 10:34 AM, Pradeep Pallan <pradeeppal...@gmail.com> wrote: > Hi Betty, > Could you post a few more screen shots of different orientations of the > electron density? > > If you see waters that are 1.6, 1.9 and 2.2 A away from the metal ion, I > would try > Mg2+ ion (trace amount of metal ions could be present in salt > solutions/buffers as an impurity) > and refine it (although 1.6Å is too close). In general, Mg2+ ions would > show an octahedral > geometry. > > > > > > On Tue, Aug 22, 2017 at 3:35 AM, <herman.schreu...@sanofi.com> wrote: > >> Dear Betty, >> >> >> >> You have very high resolution, which helps you to identify your ligand, >> but the ligand may be disordered… >> >> What I would do is to place some dummy atoms (e.g. waters) and refine and >> look if the molecule gets clearer. By scrolling the density in coot, you >> can identify the positions with the highest electron density were you >> should put your dummy atoms. >> >> >> >> Looking at your pictures, as far as is possible without the ability to >> rotate them, I would try to fit deoxyribose-phosphate, or even a complete >> nucleotide. Maybe your prep contained some unreacted nucleotides and your >> anomalous peak is phosphorus. >> >> >> >> Good luck! >> >> Herman >> >> >> >> *Von:* CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *Im Auftrag >> von *Betty Chu >> *Gesendet:* Montag, 21. August 2017 17:19 >> *An:* CCP4BB@JISCMAIL.AC.UK >> *Betreff:* [ccp4bb] Unknown positive electron density >> >> >> >> Dear ccp4bb, >> >> I am refining a 1.40 Angstrom data set for a DNA oligonucleotide. While >> the model for the DNA fits very well into the density, there is a patch of >> positive electron density in the solvent space that we are having trouble >> with. >> >> The screenshot can be viewed through this link: >> https://cbsostorage.chem.umd.edu/owncloud/index.php/s/J5cKnOpCC4vb1VC >> <https://urldefense.proofpoint.com/v2/url?u=https-3A__cbsostorage.chem.umd.edu_owncloud_index.php_s_J5cKnOpCC4vb1VC=DQMFaQ=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ=oUauCRPJVt0_cKdOakQvtJoEkS4gY9JlJmXfkd2GnlI=0XTzg6Yyi4dgz7G80vogTkpLLMZoM8fj53buMeenlJM=> >> >> In the screenshot, the yellow color is the anomalous map and a barium ion >> is fitted into density near the positive green electron density. >> >> >> The oligonucleotide was purchased from IDT. The crystallization condition >> is 15% MPD, 120 mM BaCl2, and 30 mM NaCaC pH 6.4. I have tried modelling >> Ba2+ with coordinated waters, MPD, and sodium cacodylate into the electron >> density, but none of those fit well. >> >> Any suggestions regarding the identity of this electron density is much >> appreciated. Thank you! >> >> >> >> Sincerely, >> >> Betty Chu >> >> Paukstelis Research Group >> >> Department of Chemistry and Biochemistry >> >> University of Maryland, College Park >> > > > > -- > > --- > Pradeep Pallan >
Re: [ccp4bb] Unknown positive electron density
Hi Craig, The data collection wavelength was 0.92 Angstroms. Since we observe anomalous signal for Ba at this wavelength, we would expect greater anomalous signal if As were present. There is a possibility for weak anomalous signal in this positive density, but the weak anomalous signal only shows up if I try to model a Ba in the density. Without modelling anything, there is no anomalous signal. This is what the map looks like after one round of refinement with the Ba in the density. But since there are waters that are 1.6 Angstroms, 1.9 Angstroms, and 2.2 Angstroms away from the Ba, which is smaller than the coordination distance between Ba and water, we are skeptical of the Ba being there. https://cbsostorage.chem.umd.edu/owncloud/index.php/s/O4lI2iKvRsQUoRo Thank you, Betty On Mon, Aug 21, 2017 at 5:21 PM, CRAIG A BINGMAN <cabing...@wisc.edu> wrote: > What is the data collection wavelength/energy? Would you expect > significant anomalous diffraction from As at this wavelength? > > On Aug 21, 2017, at 11:37 AM, Betty Chu <chube...@umd.edu> wrote: > > Hi Shailesh, > > When I modelled in the Barium ion with octahedrally coordinated waters and > ran the refinement, the distances from the barium to some of the waters > ended up being too close (<2.2 Angstroms). Also, the positive electron > density is connected. If the density indicated barium with coordinated > waters, would that mean there are multiple ones present in the positive > density? > > Here are more views of the connected positive density. > > https://cbsostorage.chem.umd.edu/owncloud/index.php/s/j5CgZIJe6NKgUlZ > > https://cbsostorage.chem.umd.edu/owncloud/index.php/s/YbiNFbT8AW1iHUQ > > On Mon, Aug 21, 2017 at 12:00 PM, Shailesh Tripathi <shailes...@gmail.com> > wrote: > >> Looks like Ba2+. Since it exist with coordination number 6 or above check >> what geometry water is following there (trigonal bipiramidal or so on). >> Water might also be shared by symmetry related Ba cation. >> >> >> >> Shailesh Kumar Tripathi, >> Phone: 9686289668 >> >> >> On Mon, Aug 21, 2017 at 9:17 PM, Betty Chu <chube...@umd.edu> wrote: >> >>> Yes, I have. The cacodylate ion does not fit well into the density. >>> >>> On Mon, Aug 21, 2017 at 11:44 AM, Pradeep Pallan < >>> pradeeppal...@gmail.com> wrote: >>> >>>> Did you try modelling in a cacodylate ion (CH3)2AsO2-? >>>> >>>> On Mon, Aug 21, 2017 at 10:19 AM, Betty Chu <chube...@umd.edu> wrote: >>>> >>>>> Dear ccp4bb, >>>>> >>>>> I am refining a 1.40 Angstrom data set for a DNA oligonucleotide. >>>>> While the model for the DNA fits very well into the density, there is a >>>>> patch of positive electron density in the solvent space that we are having >>>>> trouble with. >>>>> >>>>> The screenshot can be viewed through this link: >>>>> https://cbsostorage.chem.umd.edu/owncloud/index.php/s/J5cKnOpCC4vb1VC >>>>> >>>>> In the screenshot, the yellow color is the anomalous map and a barium >>>>> ion is fitted into density near the positive green electron density. >>>>> >>>>> The oligonucleotide was purchased from IDT. The crystallization >>>>> condition is 15% MPD, 120 mM BaCl2, and 30 mM NaCaC pH 6.4. I have tried >>>>> modelling Ba2+ with coordinated waters, MPD, and sodium cacodylate into >>>>> the >>>>> electron density, but none of those fit well. >>>>> >>>>> Any suggestions regarding the identity of this electron density is >>>>> much appreciated. Thank you! >>>>> >>>>> Sincerely, >>>>> >>>>> Betty Chu >>>>> Paukstelis Research Group >>>>> Department of Chemistry and Biochemistry >>>>> University of Maryland, College Park >>>>> >>>> >>>> >>>> >>>> -- >>>> >>>> --- >>>> Pradeep Pallan >>>> >>> >>> >> > >
Re: [ccp4bb] Unknown positive electron density
Hi Shailesh, When I modelled in the Barium ion with octahedrally coordinated waters and ran the refinement, the distances from the barium to some of the waters ended up being too close (<2.2 Angstroms). Also, the positive electron density is connected. If the density indicated barium with coordinated waters, would that mean there are multiple ones present in the positive density? Here are more views of the connected positive density. https://cbsostorage.chem.umd.edu/owncloud/index.php/s/j5CgZIJe6NKgUlZ https://cbsostorage.chem.umd.edu/owncloud/index.php/s/YbiNFbT8AW1iHUQ On Mon, Aug 21, 2017 at 12:00 PM, Shailesh Tripathi <shailes...@gmail.com> wrote: > Looks like Ba2+. Since it exist with coordination number 6 or above check > what geometry water is following there (trigonal bipiramidal or so on). > Water might also be shared by symmetry related Ba cation. > > > > Shailesh Kumar Tripathi, > Phone: 9686289668 > > > On Mon, Aug 21, 2017 at 9:17 PM, Betty Chu <chube...@umd.edu> wrote: > >> Yes, I have. The cacodylate ion does not fit well into the density. >> >> On Mon, Aug 21, 2017 at 11:44 AM, Pradeep Pallan <pradeeppal...@gmail.com >> > wrote: >> >>> Did you try modelling in a cacodylate ion (CH3)2AsO2-? >>> >>> On Mon, Aug 21, 2017 at 10:19 AM, Betty Chu <chube...@umd.edu> wrote: >>> >>>> Dear ccp4bb, >>>> >>>> I am refining a 1.40 Angstrom data set for a DNA oligonucleotide. While >>>> the model for the DNA fits very well into the density, there is a patch of >>>> positive electron density in the solvent space that we are having trouble >>>> with. >>>> >>>> The screenshot can be viewed through this link: >>>> https://cbsostorage.chem.umd.edu/owncloud/index.php/s/J5cKnOpCC4vb1VC >>>> >>>> In the screenshot, the yellow color is the anomalous map and a barium >>>> ion is fitted into density near the positive green electron density. >>>> >>>> The oligonucleotide was purchased from IDT. The crystallization >>>> condition is 15% MPD, 120 mM BaCl2, and 30 mM NaCaC pH 6.4. I have tried >>>> modelling Ba2+ with coordinated waters, MPD, and sodium cacodylate into the >>>> electron density, but none of those fit well. >>>> >>>> Any suggestions regarding the identity of this electron density is much >>>> appreciated. Thank you! >>>> >>>> Sincerely, >>>> >>>> Betty Chu >>>> Paukstelis Research Group >>>> Department of Chemistry and Biochemistry >>>> University of Maryland, College Park >>>> >>> >>> >>> >>> -- >>> >>> --- >>> Pradeep Pallan >>> >> >> >
Re: [ccp4bb] Unknown positive electron density
Yes, I have. The cacodylate ion does not fit well into the density. On Mon, Aug 21, 2017 at 11:44 AM, Pradeep Pallan <pradeeppal...@gmail.com> wrote: > Did you try modelling in a cacodylate ion (CH3)2AsO2-? > > On Mon, Aug 21, 2017 at 10:19 AM, Betty Chu <chube...@umd.edu> wrote: > >> Dear ccp4bb, >> >> I am refining a 1.40 Angstrom data set for a DNA oligonucleotide. While >> the model for the DNA fits very well into the density, there is a patch of >> positive electron density in the solvent space that we are having trouble >> with. >> >> The screenshot can be viewed through this link: >> https://cbsostorage.chem.umd.edu/owncloud/index.php/s/J5cKnOpCC4vb1VC >> >> In the screenshot, the yellow color is the anomalous map and a barium ion >> is fitted into density near the positive green electron density. >> >> The oligonucleotide was purchased from IDT. The crystallization condition >> is 15% MPD, 120 mM BaCl2, and 30 mM NaCaC pH 6.4. I have tried modelling >> Ba2+ with coordinated waters, MPD, and sodium cacodylate into the electron >> density, but none of those fit well. >> >> Any suggestions regarding the identity of this electron density is much >> appreciated. Thank you! >> >> Sincerely, >> >> Betty Chu >> Paukstelis Research Group >> Department of Chemistry and Biochemistry >> University of Maryland, College Park >> > > > > -- > > --- > Pradeep Pallan >
[ccp4bb] Unknown positive electron density
Dear ccp4bb, I am refining a 1.40 Angstrom data set for a DNA oligonucleotide. While the model for the DNA fits very well into the density, there is a patch of positive electron density in the solvent space that we are having trouble with. The screenshot can be viewed through this link: https://cbsostorage.chem.umd.edu/owncloud/index.php/s/J5cKnOpCC4vb1VC In the screenshot, the yellow color is the anomalous map and a barium ion is fitted into density near the positive green electron density. The oligonucleotide was purchased from IDT. The crystallization condition is 15% MPD, 120 mM BaCl2, and 30 mM NaCaC pH 6.4. I have tried modelling Ba2+ with coordinated waters, MPD, and sodium cacodylate into the electron density, but none of those fit well. Any suggestions regarding the identity of this electron density is much appreciated. Thank you! Sincerely, Betty Chu Paukstelis Research Group Department of Chemistry and Biochemistry University of Maryland, College Park
