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Alternate conformer clash problem in phenix.refine
by Aaron Oakley 20 Nov '18

20 Nov '18

Salve! I am refining a protein/ligand complex where the ligand is bound with two conformers. Conformer A is covalently bound to a cysteine residue. Conformer B is free. I have prepared an appropriate cif dictionary for the ligand including info on handling the link. The problem is that during refinement, non-bonded interactions between the two conformers appear to not be switched off and their geometry becomes distorted. Is this diagnosis correct? If so, is there a way to manually switch off interactions? With thanks, a++ Aaron Oakley Associate Professor School of Chemistry and Molecular Bioscience | Molecular Horizons | Faculty of Science, Medicine and Health University of Wollongong NSW 2522 Australia T +61 2 4221 4347 | F +61 2 4221 4287 [cid:[email protected]]

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exclude clashes for non-symmetry contacts
by Edwin Pozharski 17 Nov '18

17 Nov '18

Is there some way to make phenix.refine to ignore clashes for specific pair(s) of chains? I am dealing with a rare case of twinning where same protein molecule exists in two orientations in the same unit cell. I tried just to duplicate the molecule and set initial occupancy to 0.5, but refinement fails (as I expected) saying that there are too many close contacts. Ideally, I would like to proceed with ignoring close contacts between, say, chains A/B. I am aware that one can exclude groups of residues from symmetry clashes, but these two copies are in the same cell. I guess I could try shifting one into the next unit cell and then excluding symmetry clashes, but even assuming that will work it is a rather ugly hack. The alternative is probably to define the second orientation as an alternative conformation. Aside from aesthetic objections, I don't want to lose the ability to restrain NCS - I need two orientations to be identical structures, and I am not sure yet whether I can use NCS groups defined via alternative conformers. Cheers, Ed.

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Normalized B-factor
by Mehmet Sen 16 Nov '18

16 Nov '18

Hello, I used the Structural Comparison tool to obtain normalized B- factor values for a couple of models, each with multiple chains. Is there an efficient way to visualize the normalized B-factor values (by writing over the old B-factor values) for the models, preferably in PyMOL? Thanks, Joyce V.

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Protonation of a symmetry-related His in Zn coordination sphere
by Priit Eek 16 Nov '18

16 Nov '18

Hi, I have a structure in which one of the His-residues of a Zn coordination sphere is from a symmetry-related protein chain. I specified the symmetry in an edits file so the coordination sphere is nice and plays well in refinement. I now added hydrogens to the model, but the His from the other chain does not get protonated correctly. I assume reduce doesn’t detect the presence of the symmetry-related Zn. It protonates NE, while that is the atom that interacts with Zn, but ND should be protonated instead. What would be the best way to fix this? And another question on the same matter: is it possible to specify a LINK in PDB between these atoms? Best regards, Priit ——— Priit Eek, PhD Researcher Department of Chemistry and Biotechnology Tallinn University of Technology Tel. +372 620 4387 priit.eek(a)taltech.ee

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Postdoc Position - Berkeley
by Paul Adams 14 Nov '18

14 Nov '18

Lawrence Berkeley Laboratory Postdoctoral Researcher Job ID: 85784 Division: Molecular Biophysics & Integrated Bioimaging (Joint BioEnergy Institute) Date Opened: 11/1/2018 Summary: A postdoctoral position is immediately available to study the structure and function of macromolecular complexes and enzymes using biochemical, X-ray crystallographic and electron cryo-microscopy (cryo-EM) methods. The biological systems to be studied are: - Glycosyl transferases responsible for the synthesis of hemicelluloses in plants - Novel glycosyl hydrolases that are able to breakdown cellulosic material for biofuels production - Enzymes involved in lignin biosynthesis, degradation and valorization - Secondary metabolite and natural product enzymes This position will be part of the Structural Biology group of the Technology Division at the Joint BioEnergy Institute (JBEI) in Emeryville, California. To learn more about JBEI visit: http://www.jbei.org/ Duties: The successful candidate will have experience characterizing enzymes using biochemical techniques, and using crystallographic methods. Familiarity with cryo-EM methods would be helpful, but not essential. Robotic hardware for performing crystallization trials and imaging results are available. Crystals will be characterized and data collected using the beamline resources of the Berkeley Center for Structural Biology at the Advanced Light Source. For appropriate systems cryo-EM data will be obtained using in-house microscopes, the Bay Area Center for Cryo-EM, and other national user facilities. Qualifications: a Ph.D. in biochemistry, molecular biology, structural biology, chemistry or a related field. Demonstrated experience of scientific research using biochemistry techniques including enzymatic assays, and crystallographic methods. Experience with macromolecular crystallization techniques. Experience with cloning methods and protein purification methods would be helpful. Strong problem solving skills. Solid interpersonal skills and the ability to work in a team environment are critical. This is a full time, 2 years, postdoctoral appointment with the possibility of renewal based upon satisfactory job performance, continuing availability of funds and ongoing operational needs. You must have less than 3 years paid postdoctoral experience. Salary for Postdoctoral positions depends on years of experience post-degree. Berkeley Lab (LBNL) addresses the world’s most urgent scientific challenges by advancing sustainable energy, protecting human health, creating new materials, and revealing the origin and fate of the universe. Founded in 1931, Berkeley Lab’s scientific expertise has been recognized with 13 Nobel prizes. The University of California manages Berkeley Lab for the U.S. Department of Energy’s Office of Science. Berkeley Lab is an Equal Opportunity/Affirmative Action Employer. All qualified applicants will receive consideration for employment without regard to race, color, religion, sex, sexual orientation, gender identity, national origin, disability, age, or protected veteran status. To apply visit https://lbl.taleo.net/careersection/2/jobsearch.ftl?lang=en and search for the job number 85784 or visit: https://lbl.referrals.selectminds.com/jobs/crystallography-postdoctoral-sch… -- Paul Adams Division Director, Molecular Biophysics & Integrated Bioimaging, Lawrence Berkeley Lab Division Deputy for Biosciences, Advanced Light Source, Lawrence Berkeley Lab Adjunct Professor, Department of Bioengineering, U.C. Berkeley Vice President for Technology, the Joint BioEnergy Institute Laboratory Research Manager, ENIGMA Science Focus Area Building 33, Room 347 Building 978, Room 4126 Tel: 1-510-486-4225, Fax: 1-510-486-5909 http://cci.lbl.gov/paul Lawrence Berkeley Laboratory 1 Cyclotron Road BLDG 33R0345 Berkeley, CA 94720, USA. Executive Assistant: Louise Benvenue [ LBenvenue(a)lbl.gov ][ 1-510-495-2506 ] --

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R-free was improved, however, “Temp. fact. variance analysis” in “Validation” tool of COOT was worsened.
by Manami Torimoto 08 Nov '18

08 Nov '18

Dear all, By phenix.refine, I attempted to refine one molecule of protein. R-free was improved in certain wxc by setting the weight wxu manually, however, the results of “Temp. fact. variance analysis” in “Validation” tool of COOT was worsened. Why did such a thing happen? Thank you. Manami

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Low resolution refinement
by Ashu Sharma 06 Nov '18

06 Nov '18

Dear All, I am refining a ~7A single particle cryo-em structure of a protein. The protein has two domains and high-resolution crystal structures of individual domains are already known. I could fit these two crystal structures in the low resolution cryoem map of the full-length protein. I have two questions: 1) Is it acceptable to refine with side chains in this case? In most of the cases, with the guidane from crystal structures, side chains could be placed with reasonable confidence, particularly smaller ones. Should I restrict to poly-ala model or take the advantage of crystal structures and include side chains also? 2) When I tried to refine with side chains, longer side chains like K, R, Q, E which have weaker density tend to bend towards the backbone density. Is there a way to prevent this happening? Suggestions/comments from the community would be highly appreciated. Thanks ! Ashu

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Real-space correlation list for nucleotides?
by Boaz Shaanan 06 Nov '18

06 Nov '18

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(no subject)
by Matthew Bick 06 Nov '18

06 Nov '18

Dear Phenix BB administrator. Could you please post the following job opening to the mailing list? Thank you. Sincerely, Matthew Bick Senior Fellow Baker Laboratory for Computational Protein Design University of Washington The Institute for Protein Design, in the Department of Biochemistry at the University of Washington, currently has an outstanding opportunity for a full time X-ray Crystallographer Staff Scientist. The Institute for Protein Design is designing a new world of synthetic proteins for 21st-century challenges. PRIMARY RESPONSIBILITIES Collaborate with the Investigator and numerous researchers at the Institute for Protein Design on the X-ray crystal structure determination of de novo designed proteins. Manage the activities of research scientists and undergraduate research support with oversight of all protein-to-structure activities at the Institute for Protein Design. Design of new concepts, approaches and programs for structure determination of computationally designed proteins, of a diverse nature. Perform various specialized scientific investigative procedures in X-ray structure determination in collaborative support of numerous research projects with minimal input from Investigator. Use bio-informatics tools and pertinent laboratory and public databases, including the Protein Data Bank (PDB), and information resources to support gene-to-structure research, including deposition of atomic coordinates in the PDB for archival, sharing, and publication. Direct the activities of technical support staff in the Baker lab and at the Institute for Protein Design in collaboration with other research staff. Collaborate with other research staff, postdoctoral fellows and graduate students to select and maintain relevant protein research equipment to insure the lab meets required current and future needs. Direct the development of new protein crystallization and X-ray structure determination strategies, and ascertain if they effectively addresses needs. Manage complex multi-task projects directed at the design, production and structural characterization of numerous de novo designed proteins with higher order function. Train laboratory research staff, postdoctoral fellows and graduate students in protein X-ray crystallography, as well as wet lab equipment for protein production, structural elucidation, and biochemical characterization. Direct projects with undefined time lines and uncertain outcomes requiring analysis from multiple perspectives. Use expert knowledge and judgment to develop new methods and techniques for obtaining solutions. MINIMUM QUALIFICATIONS: Ph.D in Biochemistry, or related field, and X-ray Crystallography +5 or more years of related experience Technical proficiency, scientific creativity, collaborative ability, as well as independent thought. Advanced expertise in X-ray crystal structure determination, both computational and wet lab areas, with a full understanding of the underlying principles and concepts. May be recognized as a leader in their area of expertise. Recognized as a resource for others to use in dealing with complex protein structure determination, computational and wet lab methods, equipment, processes, or procedures. Anticipate future needs in technology support. In-depth understanding of organizational structure both within and outside the work unit, including the role of each position in the group and in other groups. Ability to develop innovative processes to achieve goals. Direct others to develop essential tasks. Review work activities to determine where new information could improve processes or move projects forward. DESIRED QUALIFICATIONS: Ph.D. degree in an appropriate field of science or technology or equivalent in education and experience AND total of six year’s research experience in protein biophysics, X-ray crystallography, computational biology and bioinformatics specifically in the X-ray crystal structure determination. Interested candidates should send a cover letter and CV to Matthew Bick at bickm(a)uw.edu.

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Speeding direct summation ?
by Clement Degut 02 Nov '18

02 Nov '18

Hi, I have (very) high resolution data, for which direct summation method seems to give significantly better map than FFT (i.e. visible hydrogen vs not). My main problem is that refinement got from quite slow, to barely not really possible to handle slow. I have 2 question from that : Am I looking at an artifact ? Meaning should at this resolution (0.84A for CC1/2 at 30% and good overall statistics) see extreme difference between map quality with direct summation vs FFT ? And if yes, can I speed the process compiling phenix with openMP on our cluster ? Or shoul I just become patient ? Many thanks Clément

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