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Have you performed MS on the sample/crystals to determine the mass of the O-linked species? If there is partial occupancies as Herman suggest you should see two species in the MS data (if they both fly). Dan Daniel A Bonsor PhD. Sundberg Lab Institute of Human Virology University of Maryland, Baltimore 725 W Lombard Street N370 Baltimore Maryland MD 21201 Tel: (410) 706-7457 From: CCP4 bulletin board [mailto:[log in to unmask]] On Behalf Of [log in to unmask] Sent: Tuesday, September 13, 2016 10:01 AM To: [log in to unmask] Subject: [ccp4bb] AW: [ccp4bb] Unknown density-reg Dear Gayathri, You may have incomplete modification. You could try to make alternate Tyr and Tys/Ptr and play around with occupancies. Herman Von: CCP4 bulletin board [mailto:[log in to unmask]] Im Auftrag von Chellam Gayathri.S bt11d002 Gesendet: Dienstag, 13. September 2016 15:40 An: [log in to unmask]<mailto:[log in to unmask]> Betreff: Re: [ccp4bb] Unknown density-reg Dear Gyanendra Kumar, I have tried modelling both TYS ( sulfo tyrosine) and PTR ( phospho tyrosine) in the density but with negative peaks in the Fo-Fc maps. I would like to know if there could be any other modifications of tyrosine. Thank you, Gayathri Subash On Tue, Sep 13, 2016 at 6:49 PM, Gyanendra Kumar <[log in to unmask]<mailto:[log in to unmask]>> wrote: Have you tried to fit O-phosphate or O-sulfate in this density? On Mon, Sep 12, 2016 at 12:36 AM, Chellam Gayathri.S bt11d002 <[log in to unmask]<mailto:[log in to unmask]>> wrote: Dear All, I am facing difficulty in interpreting an unknown density on a surface tyrosine residue. It appears like an O-modified tyrosine. Two lysines and one histidine are other residues in its proximity. My storage buffer contains 20mM HEPES, 200mM NaCl and 2mM DTT. The protein was purified with an imidazole gradient and buffer exchanged with the storage buffer. Crystallization buffers include 2.0 M Ammonium sulfate, 0.2 M Na K tartarate and 0.1 M Na citrate, pH 5.6. 10% Glycerol and 3.2 M Ammonium sulfate were used as cryo. The density was too small for tartrate, while the Fo-Fc map showed some positive density around a fully occupied glycerol. I was not able to model any of the other buffer components in it. Attached are the links to the images of the structure. Blue is 2Fo-Fc, contoured to 1σ and green is Fo-Fc contoured to 3σ. The structure was solved to 2.0 Å resolution. The density appears in all the crystals solved in this condition. I would like to know if anyone has come across such a density. TIA for your suggestions. https://drive.google.com/open?id=0B6r9xAuEQwQKM0EtZlNBcjFNLWs https://drive.google.com/open?id=0B6r9xAuEQwQKT3B3Zmktai1mc1k https://drive.google.com/file/d/0B6r9xAuEQwQKejc2OUQyeldhWFk/view?usp=sharing -- Gayathri Subash Research Scholar, Lab No: 309, Structural Biology Lab, IIT Madras, Chennai-600036 -- Gyanendra Kumar, PhD St. Jude Children's Research Hospital, Department of Structural Biology, 262, Danny Thomas Place, MS-311 Memphis, TN 38105 Phone: 901-595-3839 Cell: 631-875-9189 https://www.linkedin.com/in/gyanendrakumar https://scholar.google.com/gyanendrakumar<https://scholar.google.com/citations?user=lnil0IQAAAAJ&hl=en> https://www.researchgate.net/profile/Gyanendra_Kumar3 http://stjude.academia.edu/GyanendraKumar -------------------------------------------------------