- phenixbb - phenix-online.org

Phenix version 1.19 released
by Paul Adams 11 Jan '21

11 Jan '21

The Phenix developers are pleased to announce that version 1.19 of Phenix is now available (build 1.18.2-3874). Binary installers for Linux, Mac OSX, and Windows platforms, and the source installer, are available at the download site: http://phenix-online.org/download/ Please note that this recent publication should now be used to cite the use of Phenix: Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix. Liebschner D, Afonine PV, Baker ML, Bunkóczi G, Chen VB, Croll TI, Hintze B, Hung LW, Jain S, McCoy AJ, Moriarty NW, Oeffner RD, Poon BK, Prisant MG, Read RJ, Richardson JS, Richardson DC, Sammito MD, Sobolev OV, Stockwell DH, Terwilliger TC, Urzhumtsev AG, Videau LL, Williams CJ, Adams PD: Acta Cryst. (2019). D75, 861-877. https://doi.org/10.1107/S2059798319011471 Highlights for the 1.19 version of Phenix: - phenix.real_space_refine: - Improved rotamer fitting (use multiprocessing, in case of NCS constraints work on one copy only and propagate changes to all related copies, various performance improvements and bug fixes) - Improved map/restraints weight calculation - Morphing can now use multiprocessing (nproc) - phenix.map_to_model and phenix.trace_and_build - Improved high-resolution model-building including detection of insertions and deletions - New methods - phenix.local_resolution: calculates a local resolution map - phenix.local_aniso_sharpen: optimizes a map taking into account local resolution-dependence and anisotropy of the map and its errors - New scripting tools - High-level scriptable Python tools are now available for map and model analyses and manipulation as well as for model-building - Restraints - Adjusting the "positions" of atom names in (pseudo-)symmetric amino acid side chains is now the default - Improved restraints for ARG allows more flexibility of the CD atom - Bug fixes - Chains with modified amino or nucleic acids can be aligned - Fixed handling of modified amino/nucleic acids in sequence alignment For a full list of changes see: http://www.phenix-online.org/documentation/CHANGES Please note that this new publication should be used to cite the use of Phenix: Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix. Liebschner D, Afonine PV, Baker ML, Bunkóczi G, Chen VB, Croll TI, Hintze B, Hung LW, Jain S, McCoy AJ, Moriarty NW, Oeffner RD, Poon BK, Prisant MG, Read RJ, Richardson JS, Richardson DC, Sammito MD, Sobolev OV, Stockwell DH, Terwilliger TC, Urzhumtsev AG, Videau LL, Williams CJ, Adams PD: Acta Cryst. (2019). D75, 861-877. https://doi.org/10.1107/S2059798319011471 Full documentation is available here: http://www.phenix-online.org/documentation/ There is a Phenix bulletin board: http://www.phenix-online.org/mailman/listinfo/phenixbb/ Please consult the installer README file or online documentation for installation instructions. Direct questions and problem reports to the bulletin board or: help(a)phenix-online.org and bugs(a)phenix-online.org Commercial users interested in obtaining access to Phenix should visit the Phenix website for information about the Phenix Industrial Consortium. The development of Phenix is principally funded by the National Institute of General Medical Sciences (NIH) under grant P01-GM063210. We also acknowledge the generous support of the members of the Phenix Industrial Consortium. -- Paul Adams Division Director, Molecular Biophysics & Integrated Bioimaging, LBL (http://biosciences.lbl.gov/divisions/mbib) Principal Investigator, Computational Crystallography Initiative, LBL (http://cci.lbl.gov) Vice President for Technology, the Joint BioEnergy Institute (http://www.jbei.org) Principal Investigator, ALS-ENABLE, Advanced Light Source (http://als-enable.lbl.gov) Division Deputy for Biosciences, Advanced Light Source (https://als.lbl.gov) Laboratory Research Manager, ENIGMA Science Focus Area (http://enigma.lbl.gov) Adjunct Professor, Department of Bioengineering, U.C. Berkeley (http://bioeng.berkeley.edu) Adjunct Professor, Comparative Biochemistry, U.C. Berkeley (http://compbiochem.berkeley.edu) Building 33, Room 250 Building 978, Room 4126 Building 977, Room 180C Tel: 1-510-486-4225 http://cci.lbl.gov/paul Lawrence Berkeley Laboratory 1 Cyclotron Road BLDG 33R0345 Berkeley, CA 94720, USA. Executive Assistant: Ashley Dawn [ ashleydawn(a)lbl.gov ][ 1-510-486-5455 ] --

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Post-doc vacancy in cryoem of viruses
by Hasan, Syed Saif 11 Jan '21

11 Jan '21

Dear Colleagues, A vacancy for a post-doctoral researcher is available immediately in my lab in the University of Maryland Center for Biomolecular Therapeutics located on the campus of Institute for Bioscience and Biotechnology Research (IBBR; https://www.ibbr.umd.edu/profiles/s-saif-hasan<https://nam11.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.ibbr.…>) in Rockville MD. We use high-resolution single particle cryoEM and X-ray crystallography to investigate how viruses utilize structural similarities with host proteins to hijack inter-organelle communication. Prior experience in insect cell culture for recombinant protein production is desirable. Expertise in cryoEM or X-ray crystallography will be advantageous. Candidates who have recently completed their PhD are encouraged to apply. IBBR is located 20 miles north-west of Washington DC. In IBBR, you will have in-house access to a 200 kV Talos Arctica with a Falcon 3EC DED and a 200 kV Glacios with a K3 DED and a Volta Phase Plate (VPP). We have a Vitrobot Mark IV and a Cp3 cryo-plunger for sample preparation. We have off-site access to a Titan Krios microscope equipped with DED and VPP. For X-ray crystallography, we have access to a Mosquito crystallization robot. Data collection is usually performed at synchrotrons. Our research is highly collaborative and is supported by wonderful colleagues who help us out with their expertise in NMR spectroscopy, lipidomics, HDX-MS, MD simulations, computer assisted drug design, and cellular imaging. Interested candidates should send their CV and a cover letter to me at sshasan(a)som.umaryland.edu ************************************************************************************************ The University of Maryland, Baltimore is an Equal Opportunity, Affirmative Action employer. Minorities, women, individuals with disabilities and protected veterans are encouraged to apply. ************************************************************************************************ Saif S. Saif Hasan, PhD Assistant Professor Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine 108 N. Greene Street, Baltimore MD 21201 https://www.medschool.umaryland.edu/profiles/Hasan-S-Saif-Hasan-Syed-Saif/ Associate Member Program in Oncology, University of Maryland Marlene and Stewart Greenebaum Comprehensive Cancer Center (UMGCCC) 22 S. Greene Street, Baltimore MD 21201 Office and Laboratory Address: Center for Biomolecular Therapeutics, Institute for Bioscience and Biotechnology Research (IBBR) 9600 Gudelsky Drive, Rockville MD 20850 https://www.ibbr.umd.edu/profiles/s-saif-hasan Phone: 240-314-6396 Fax: 240-314-6225

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Senior Technician Position at the Biochemistry and Structural Biology Unit - IEO Milano
by Sebastiano Pasqualato 11 Jan '21

11 Jan '21

Dear all, a position is open at the Biochemistry and Structural Biology Unit of the European Institute of Oncology in Milan. The Unit (https://www.research.ieo.it/research-and-technology/technological-units/bio… <https://www.research.ieo.it/research-and-technology/technological-units/bio…>) is one of the technological platforms of the Department of Experimental Oncology of the European Institute of Oncology (www.research.ieo.it <http://www.ieo0researh.it/>), a comprehensive cancer centre located in the south area of Milan. We provide services and support to the department in the fields of Biochemistry and Structural Biology and collaborate both with scientific groups of the department and external ones to foster the structural biology aspects of their projects. The ideal candidate has a Master/PhD degree in Chemistry, Biotechnology or Biology, with strong interest in biochemistry and structural biology. We are looking for an enthusiastic, motivated person, with good organisational and relational skills, and a good knowledge of English. She/he will take care of assisting scientists collaborating with the Unit in macromolecular samples purification, characterisation and crystallisation, managing sample shipment to synchrotrons, supervising instrumentation usage and maintenance. The Unit is setting up a Cryo-EM lab with an in-house VitroBot blotting device and external Electron Microscopy access, which the candidate is expected to manage. Experience in setting up grids and Cryo-EM data collection will be a distinguishing skill. Potential applicants are encouraged to contact Dr. Sebastiano Pasqualato, sending a brief description of interests, and a CV with at least one referee contact to sebastiano.pasqualato(a)ieo.it <mailto:[email protected]>. Best regards, Sebastiano -- Sebastiano Pasqualato, PhD Biochemistry and Structural Biology Unit Department of Experimental Oncology European Institute of Oncology IFOM-IEO Campus via Adamello, 16 20139 - Milano Italy tel +39 02 9437 5167 fax +39 02 9437 5990 web: https://www.research.ieo.it/research-and-technology/technological-units/bio…

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Postdoc position available on membrane proteins at Aarhus University
by Joseph Lyons 11 Jan '21

11 Jan '21

Dear Colleagues, Please pass this along to anyone who might be interested. Thanks! Joe online posting: http://tiny.cc/y1y7tz We are looking for a highly skilled and motivated postdoc with an interest in working on structural studies of lipid transporting membrane transporters by cryo-EM. The applicant should preferably have a proven track record in the area of structural and/or functional analysis of membrane proteins. The position will be open from Spring 2021, but starting date is negotiable. Funding is available for at least 2 years of employment, with an extension possible for up to a total maximum of 3 years. The position: The position aims to strengthen ongoing activities in the newly established Lyons Lab at the Department of Molecular Biology and Genetics, Aarhus University, Denmark. The laboratory's interest is the interplay between structure and function of transmembrane transport processes with a focus on lipid transport systems. The lab houses facilities for mammalian protein production, purification, cellular, biochemical and biophysical characterization and data processing and has access to the state-of-the-art Cryo-EM facility based at iNANO (https://inano.au.dk/research/research-platforms/inano-cryo-em-facility/). The group is part of the Section of Structural Biology at Aarhus University. The project is within the framework of a highly prestigious and international collaborative research program ASAP (Aligning Science Across Parkinson’s), the team will explore the structure, function and regulation of the human glucosylceramide transporters (ATP10) (https://parkinsonsroadmap.org/research-network/biology-of-pd-associated-gen…). These molecular insights may pave the way for a drug discovery strategy for Parkinson’s disease. The candidate: A successful candidate has a relevant Ph.D. degree and a solid and documented background in structural biology, biochemistry and/or biophysics. Experience with membrane protein expression and purification and/or functional characterization is favored, and the candidate must demonstrate an ability and interest to work with membrane proteins with a structural aim. Applicants should be ambitious, show strong collaborative skills, and be able to take initiatives and responsibility within the work environment. The successful candidate is offered - a full-time appointment (100%) as a post doc for a period of 3 years with the possibility of an extension. The successful candidate is encouraged to initiate and participate in grant proposals to expand the research line and ensure continuation after the project end - Participation in a vibrant international research network ASAP with focus on collaboration and open science, offering excellent networking opportunities. - access to a well-developed research infrastructure. - a research climate inviting lively, open and critical discussion within and across different fields of research. - a working environment with teamwork, close working relations, network activities among young scientists and social activities. - a workplace characterized by professionalism, equality and a healthy work-life balance. The city: In Aarhus, you have easy access to beautiful nature, an exciting culture and city life as well as a safe environment for children - a great place for the whole family. The city of Aarhus has everything you need: exciting national and international jobs, delightful residential areas, a rich cultural life, and beautiful surrounding landscape of woods and coastline that make Aarhus a wonderful place to live and work. Deadline: All applications must be made online (http://tiny.cc/y1y7tz) and received by 7th Feb. 2021.

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About Restrain Files for ligands
by Sharan Karade 08 Jan '21

08 Jan '21

Hello all, I hope everybody is doing well. Happy New Year 2021!!! I am stuck with some unusual behavior of restrain files generated by Phenix.ReadySet. I crystallized the protein with ligands which have double bonds between the atoms. As I see clear electron density for the ligand, I drew a ligand molecule in Coot (Ligand builder) and fitted in electron density very nicely. Until here, I can see the double bonds in the modelled ligand. I generated the restrain files using Phenix.ReadySet to further proceed for refinement. Unfortunately, after refinement the double bonds were converted to single bonds. I tried to generate new restrain files and refined the model, resulting in similar output. But, fortunately in one case, the double bonds were retained after refinement also. I tried to repeat with other ligands, where I was unsuccessful to retain the double bonds. please help me. Thank you so much. Regards Sharan -- Sharan Karade Postdoc-fellow IBBR-UMD, 9600 Gudelsky Dr, Rockville Maryland 20850

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Modeling sugars with Coot and Phenix
by Emilia C. Arturo (Emily) 07 Jan '21

07 Jan '21

Hello All, I have several questions regarding sugar modeling. These are questions for both Coot (0.9.4-pre/CCPEM) and Phenix (1.18.2-3874) usage (on a Mac) for modeling as correctly as possible several N-linked glycosylation (i.e. not ligand) sugars within a protein, and about the approach as it pertains to higher resolution X-ray maps (better than 2A) or to lower resolution EM maps (~3-4A). I am new to the world of glycoproteins and cryoEM, not new to model building or X-ray crystallography, and would welcome any advice or insight. For relatively low resolution maps, where sugar moieties cannot be placed unambiguously, in theory (say Agirre (2017) and Emsley and Crispin (2018)) pyranose placement within the map should be quite highly restrained by ideal geometry for the specific sugar expected from prior knowledge. In practice, however, it’s not entirely clear how to maintain these restraints on pyranose geometry and conformation during manual building in Coot, or how to apply any special restraints in Phenix. The Glyco module of Coot does indeed place fine looking Asn-NAGs after a few iterations of manual placing and refining to account for the poor resolution of the acetyl group, just as discussed by Emsley and Crispin (2018). However, the validation report that is obtained from the OneDep system at the rcsb web server for the resulting structure shows many outliers in bonds and angles within the NAG or between the NAG and protein Asn residue. In Coot I have experimented with various restraints (in the R/RC menu), including decreasing the Refinement weight/weight matrix to zero, or some small non-zero number, thinking this places less weight on the map signal and more on the ideal values within Coot’s libraries. I have tried a variety of refinement restraint combinations in Coot to address individual bonds and angles outliers within the sugar, but the rcsb validation report still finds departures from ideal geometry. Meanwhile, in Phenix I don’t do anything specific for the refinement of glycosylation sugars (should I?) other than apply the appropriate restraints for any low resolution EM map; are there specific restraints files I ought to be using within Phenix real space refine jobs for N-linked sugars? Then I noticed that loads of existing PDB entries containing N-linked glycans contain similar outliers in pyranose geometry. So this prompted me to wonder whether we’re not doing things right, that in practice we’re not correctly following what’s advised in theory, or whether this is just the state of model building currently. Could someone more experienced than am I on sugar building please comment here on how exactly to get the sugars better built? e.g. what buttons to press and why, in Phenix and/or in Coot. Or is this a tolerable situation where sugar geometries differ from ideal even at low resolution? Is it perhaps a matter of different targets being used for PDB validation by the rcsb versus by Coot? It turns out that regarding higher resolution maps I am also unsure of exactly how to apply restraints within Coot. For example, using PDB ID 5VAA and its map with a reported resolution of 1.55A, I centered on Asn297 and its associated glycosylation sugars. I experimented with refinement strategies (i.e. the settings of R/RC), thinking that the best approach would be little restraint on ideal geometry and higher weight on the map signal. But no matter what combination of restraints I used, I was unable to keep the model locally from distorting; i.e. the deposited model versus map looks great, but within Coot I find a hundred ways to destroy it. I must be doing something wrong. Any advice or references you’d recommend would be greatly appreciated. Thanks. -- "Study as if you were going to live forever; live as if you were going to die tomorrow." - Maria Mitchell "I'm not afraid of storms for I'm learning to sail my ship." - Louisa May Alcott

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post-doc position at KU Leuven, ligand-gated ion channels
by Chris Ulens 05 Jan '21

05 Jan '21

A post-doctoral position is available at the Laboratory of Prof. Chris Ulens, KU Leuven, Belgium (http://www.xtal.be <http://www.xtal.be/>). The research in the Laboratory of Structural Neurobiology is aimed at understanding the molecular mechanism of ligand-gated ion channels. Our methods rely on X-ray crystallography and cryo-EM in combination with electrophysiological recordings and fluorescence-based techniques. Our laboratory hosts an integrated macromolecular crystallization facility with a Mosquito crystallization robot and two RockImagers. We have access to a 200 kV Glacios microscope on campus as well as 300 kV JEOL CryoARM300 at the nearby EM facility in Brussels. Current projects are situated at the crossroads of structural elucidation and drug discovery of different pentameric ligand-gated ion channels. We would like to recruit a post-doc with prior experience in cryo-EM or X-ray crystallography of membrane proteins. The successful candidate: - easily communicates in an open verbal manner and has a team player spirit - has an ambitious work ethic - critically contributes to writing of manuscripts and grant proposals - makes efforts to obtain independent funding - meets intermediate deadlines and brings projects to a successful end - fuels internal discussions of recent publications in the field - sets an example to younger PhD students in the lab - identifies directions for future scientific research The position is open immediately and applications will be accepted until the position is filled. Applicants are requested to submit a CV with publication list and two references. E-mail to chris.ulens(a)kuleuven.be <mailto:[email protected]> Selection of recent publications: A lipid site shapes the agonist response of a pentameric ligand-gated ion channel. Hénault CM, Govaerts C, Spurny R, Brams M, Estrada-Mondragon A, Lynch J, Bertrand D, Pardon E, Evans GL, Woods K, Elberson BW, Cuello LG, Brannigan G, Nury H, Steyaert J, Baenziger JE, Ulens C. Nat Chem Biol. 2019;15(12):1156-1164. doi: 10.1038/s41589-019-0369-4. Modulation of the Erwinia ligand-gated ion channel (ELIC) and the 5-HT3 receptor via a common vestibule site. Brams M, Govaerts C, Kambara K, Price KL, Spurny R, Gharpure A, Pardon E, Evans GL, Bertrand D, Lummis SC, Hibbs RE, Steyaert J, Ulens C. Elife. 2020;9:e51511. doi: 10.7554/eLife.51511. The structural basis for an on-off switch controlling Gβγ-mediated inhibition of TRPM3 channels. Behrendt M, Gruss F, Enzeroth R, Dembla S, Zhao S, Crassous PA, Mohr F, Nys M, Louros N, Gallardo R, Zorzini V, Wagner D, Economou A, Rousseau F, Schymkowitz J, Philipp SE, Rohacs T, Ulens C, Oberwinkler J. Proc Natl Acad Sci U S A. 2020;117(46):29090-29100. doi: 10.1073/pnas.2001177117.

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Bad geometry for DNA after Real-Space-Refine
by Jobichen Chacko 05 Jan '21

05 Jan '21

Dear BB, I am using phenix real space refinement for refining a cryo-EM model of protein-DNA complex. However after refinement the structure of the dna strands are twisted in a very bad geometry compared to the initial input model. I am using version 1.8 for this project, also tried couple of other versions as well, but the same problem happened. How to rectify this issue? Thank you. Jobi

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Variation in bonds after loading restraints
by Vatsal Purohit 01 Jan '21

01 Jan '21

Hello, I’ve been having issues with obtaining consistent number of bonds after generating .cif files using eLBOW. To generate the .cif file for an NADH molecule on eLBOW, I’ve been using a SMILE string generated using Chemdraw and a template .pdb file that was generated after using LigFit with a ligand .pdb model (also initially generated using a SMILE string). However, I find that when I load the restraints the bonds in the molecule only show correctly on the older version of coot (0.8.9.2). The double bonds on the adenine ring don’t appear in the latest version of coot (0.9.3) and the double bonds on the molecule don’t show up at all when I load the .cif file on REEL. Is there a reason why they could be inconsistent? Could I be viewing them incorrectly on REEL? Any thoughts on this would be appreciated! I’ve attached the SMILE string, template .pdb file and final .cif file in this email for your reference. Regards, Vatsal PhD Candidate, Stauffacher Lab, Dept. of Biology, Purdue University PULSe-Biophysics and Structural Biology training group vpurohi(a)purdue.edu<mailto:[email protected]> | 346-719-9409

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Calculating the transformation matrix of density map alignment
by Ahmad Khalifa 23 Dec '20

23 Dec '20

I have two density maps that I'm trying to align, or more precisely, calculate the transformation matrix (rotation and translation) of their alignment (moving one map and aligning it with the other map). Is there a way to accomplish that using phenix or any other tool? Thanks a lot and happy holidays.

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