Map challenge: Analysis using a pair comparison method based on Fourier shell correlation

This document presents the analysis performed over the Map Challenge dataset using a new algorithm which we refer to as Pair Comparison Method. The new algorithm, which is described in detail in the text, is able to sort reconstructions based on a figure of merit and assigns a level of significance to the sorting. That is, it shows how likely the sorting is due to chance or if it reflects real differencesThe authors would like to acknowledge economical support from: The Spanish Ministry of Economy and Competitiveness through Grants BIO2013-44647-R, BIO2016-76400-R(AEI/FEDER, UE) and AEI/FEDER BFU 2016 74868P, the Comunidad Autónoma de Madrid through Grant: S2017/BMD-3817, European Union (EU) and Horizon 2020 through grant CORBEL (INFRADEV-1-2014-1, Proposal: 654248). This work used the EGI Infrastructure and is co-funded by the EGI-Engage project (Horizon 2020) under Grant No. 654142. European Union (EU) and Horizon 2020 through grant West-Life (EINFRA-2015-1, Proposal: 675858) European Union (EU) and Horizon 2020 through grant Elixir - EXCELERATE (INFRADEV-3-2015, Proposal: 676559) European Union (EU) and Horizon 2020 through grant iNEXT (INFRAIA-1–2014-2015, Proposal: 653706). The authors acknowledge the support and the use of resources of Instruct, a Landmark ESFRI projec

Particle alignment reliability in single particle electron cryomicroscopy: A general approach

Electron Microscopy is reaching new capabilities thanks to the combined effect of new technologies and new image processing methods. However, the reconstruction process is still complex, requiring many steps and elaborated optimization procedures. Therefore, the possibility to reach a wrong structure exists, justifying the need of robust statistical tests. In this work, we present a conceptually simple alignment test, which does not require tilt-pair images, to evaluate the alignment consistency between a set of projection images with respect to a given 3D density map. We test the approach on a number of problems in 3DEM, especially the ranking and evaluation of initial 3D volumes and high resolution 3D maps, where we show its usefulness in providing an objective evaluation for maps that have recently been subject to a strong controversy in the field. Additionally, this alignment statistical test can be linked to the early stages of structure solving of new complexes, streamlining the whole process.The authors would like to acknowledge economical support from the Spanish Ministry of Economy and Competitiveness through grants AIC-A-2011-0638 and BIO2013-44647-R, the Comunidad de Madrid through grant CAM (S2010/BMD-2305), and from "Comfuturo" grant funded by the Fundacion General CSIC. C.O.S. Sorzano is recipient of a Ramon y Cajal fellowship. We acknowledge discussions with Sriram Subramanian on validation approaches related to the HIV trimer structure

Transfer function restoration in 3D electron microscopy via iterative data refinement

Three-dimensional electron microscopy (3D-EM) is a powerful tool for visualizing complex biological systems. As with any other imaging device, the electron microscope introduces a transfer function (called in this field the contrast transfer function, CTF) into the image acquisition process that modulates the various frequencies of the signal. Thus, the 3D reconstructions performed with these CTF-affected projections are also affected by an implicit 3D transfer function. For high-resolution electron microscopy, the effect of the CTF is quite dramatic and limits severely the achievable resolution. In this work we make use of the iterative data refinement (IDR) technique to ameliorate the effect of the CTF. It is demonstrated that the approach can be successfully applied to noisy data.Partial support is acknowledged to the Comisión Interministerial de Ciencia y Tecnología of Spain through projects BIO98-0761 and BIO2001-1237 and to National Institutes of Health through grant HL70472. The work of Y. Censor was done in part at the Center for Computational Mathematics and Scientific Computation (CCMSC) at the University of Haifa and supported by Research Grant 592/00 from the Israel Science Foundation founded by the Israel Academy of Sciences and Humanities

Optimization problems in electron microscopy of single particles

The final publication is available at Springer via http://dx.doi.org/10.1007/s10479-006-0078-8Electron Microscopy is a valuable tool for the elucidation of the three-dimensional structure of macromolecular complexes. Knowledge about the macromolecular structure provides important information about its function and how it is carried out. This work addresses the issue of three-dimensional reconstruction of biological macromolecules from electron microscopy images. In particular, it focuses on a methodology known as “single-particles” and makes a thorough review of all those steps that can be expressed as an optimization problem. In spite of important advances in recent years, there are still unresolved challenges in the field that offer an excellent testbed for new and more powerful optimization techniques.We acknowledge partial support from the “Comunidad Autónoma de Madrid” through grants CAM-07B-0032-2002, GR/SAL/0653/2004 and GR/SAL/0342/2004, the “Comisión Interministerial de Ciencia yTecnologia” of Spain through grants BIO2001-1237, BIO2001-4253-E, BIO2001-4339-E, BIO2002- 10855-E, BFU2004-00217/BMC, the Spanish FIS grant (G03/185), the European Union through grants QLK2- 2000-00634, QLRI-2000-31237, QLRT-2000-0136, QLRI-2001-00015, FP6-502828 and the NIH through grant HL70472. Alberto Pascual and Roberto Marabini acknowledge support by the Spanish Ramon y Cajal Program

On the development of three new tools for organizing and sharing information in three-dimensional electron microscopy

This work was funded by the Spanish Ministerio de Economía y Competividad through grants BFU2009-09331, BIO2010-16566, ACI2009-1022, ACI2010-1088 and AIC-A- 2011-0638, by the Comunidad Autonoma de Madrid through grant S2010/BMD-2305, by NFS grant No. 1114901 and by the Spanish National Institute of Bioinformatics (a project funded by the Instituto de Salud Carlos III). This work was conducted using the Protégé resource, which is supported by grant LM007885 from the United States National Library of Medicine. COSS is a Ramón y Cajal researcher financed by the European Social Fund and the Ministerio de Economía y Competitividad. JV is a Juan de la Cierva Postdoctoral Fellow (JCI-2011-10185). This work was funded by Instruct, which is part of the European Strategy Forum on Research Infrastructures (ESFRI) and is supported by national member subscriptions

Consistent and elastic registration of histological sections using vector-spline regularization

The final publication is available at Springer via http://dx.doi.org/10.1007/11889762_8Revised Papers on Second International ECCV Workshop, CVAMIA 2006 Graz, Austria, May 12, 2006Here we present a new image registration algorithm for the alignment of histological sections that combines the ideas of B-spline based elastic registration and consistent image registration, to allow simultaneous registration of images in two directions (direct and inverse). In principle, deformations based on B-splines are not invertible. The consistency term overcomes this limitation and allows registration of two images in a completely symmetric way. This extension of the elastic registration method simplifies the search for the optimum deformation and allows registering with no information about landmarks or deformation regularization. This approach can also be used as the first step to solve the problem of group-wise registration.Ignacio Arganda-Carreras is being supported by a predoctoral FPI-CAM fellow- ship since October 2003. Carlos Ortiz-de-Solorzano is supported by a Ramon y Cajal (Spanish Ministry of Education and Science ryc-2004-002353) and a Marie Curie International Reintegration Grant (FP6-518688). Jan Kybic was sponsored by the Czech Ministery of Education under project number MSM210000012. Par- tial support is acknowledged to Comunidad de Madrid through grant GR/SAL/0234, to Instituto de Salud Carlos III-Fondo de Investigaciones Sanitarias (FIS) through the IM3 Network and grant 040683 and to the Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica (I+D+I)

Cryo-EM structure of enteric adenovirus HAdV-F41 highlights structural variations among human adenoviruses

Enteric adenoviruses, one of the main causes of viral gastroenteritis in the world, must withstand the harsh conditions found in the gut. This requirement suggests that capsid stability must be different from that of other adenoviruses. We report the 4-Å-resolution structure of a human enteric adenovirus, HAdV-F41, and compare it with that of other adenoviruses with respiratory (HAdV-C5) and ocular (HAdV-D26) tropisms. While the overall structures of hexon, penton base, and internal minor coat proteins IIIa and VIII are conserved, we observe partially ordered elements reinforcing the vertex region, which suggests their role in enhancing the physicochemical capsid stability of HAdV-F41. Unexpectedly, we find an organization of the external minor coat protein IX different from all previously characterized human and nonhuman mastadenoviruses. Knowledge of the structure of enteric adenoviruses provides a starting point for the design of vectors suitable for oral delivery or intestinal targetingThis work was supported by grants PID2019-104098GB-I00/AEI/10.13039/501100011033 and BFU2016-74868-P, cofunded by the Spanish State Research Agency and the European Regional Development Fund; BFU2013-41249-P and BIO2015-68990-REDT (the Spanish Adenovirus Network, AdenoNet) from the Spanish Ministry of Economy, Industry, and Competitiveness; and the Agencia Estatal CSIC (2019AEP045) to C.S.M. The CNB-CSIC is further supported by a Severo Ochoa Excellence grant (SEV 2017-0712). Work in M.B’s. lab was supported by grant 194562-08 from the Natural Sciences and Engineering Research Council of is a recipient of a Juan de la Cierva postdoctoral contract funded by the Spanish State Research Agency. M.P.-I. holds a predoctoral contract from La Caixa Foundation (ID 100010434), under agreement LCF/BQ/SO16/52270032. Access to CEITEC was supported by iNEXT, project number 653706, funded by the Horizon 2020 Programme of the European Union. The CEITEC Cryo-electron Microscopy and Tomography core facility is supported by MEYS CR (LM2018127

MRC2014: Extensions to the MRC format header for electron cryo-microscopy and tomography

Open Access funded by Medical Research CouncilThe MRC binary file format is widely used in the three-dimensional electron microscopy field for storing image and volume data. Files contain a header which describes the kind of data held, together with other important metadata. In response to advances in electron microscopy techniques, a number of variants to the file format have emerged which contain useful additional data, but which limit interoperability between different software packages. Following extensive discussions, the authors, who represent leading software packages in the field, propose a set of extensions to the MRC format standard designed to accommodate these variants, while restoring interoperability. The MRC format is equivalent to the map format used in the CCP4 suite for macromolecular crystallography, and the proposal also maintains interoperability with crystallography software. This Technical Note describes the proposed extensions, and serves as a reference for the standard.We thank Chris Booth and Steffen Meyer from Gatan Inc. for clarifying the format definition used by Digital Micrograph. Acknowledgement for support from National Institute of Health, USA includes: NIGMS grant P41GM103310 (AC and SD), NIBIB grant 5R01-EB005027 (DM), and R01GM080139 (SJL). RH and MW would like to thank the UK Medical Research Council for the award of Partnership Grant MR/J000825/1 to support the establishment of CCP-EM. RH and JS are also supported by MRC grant U105184322

FSC-Q: a CryoEM map-to-atomic model quality validation based on the local Fourier shell correlation

In recent years, advances in cryoEM have dramatically increased the resolution of reconstructions and, with it, the number of solved atomic models. It is widely accepted that the quality of cryoEM maps varies locally; therefore, the evaluation of the maps-derived structural models must be done locally as well. In this article, a method for the local analysis of the map-to-model fit is presented. The algorithm uses a comparison of two local resolution maps. The first is the local FSC (Fourier shell correlation) between the full map and the model, while the second is calculated between the half maps normally used in typical single particle analysis workflows. We call the quality measure “FSC-Q”, and it is a quantitative estimation of how much of the model is supported by the signal content of the map. Furthermore, we show that FSC-Q may be helpful to detect overfitting. It can be used to complement other methods, such as the Q-score method that estimates the resolvability of atomsWe thank Prof. David Veesler for providing us the half maps of the spike glycoprotein of SARS-CoV-2. The authors would like to acknowledge financial support from: the Comunidad de Madrid through grant CAM (S2017/BMD-3817), the Spanish National Research Council (PIE/COVID-19 number 202020E079), the Spanish Ministry of Economy and Competitiveness through grants SEV 2017-0712, PID2019-104757RB-I00/AEI/10.13039/501100011033, the Instituto de Salud Carlos III through grant PT17/0009/0010 (ISCIII-GEFI/ERDF-). Instruct-ULTRA (Grant 731005), an EU H2020 project to further develop the services of Instruct-ERIC. UE H2020 grant HighResCells (ERC-2018-SyG, Proposal: 810057). This work was supported by the Intramural Research Program of the National Institute for Arthritis, musculoskeletal, and Skin Diseases, NIH. The authors acknowledge the support and the use of resources of Instruct, a Landmark ESFRI projec

Cryo-EM and the elucidation of new macromolecular structures: Random Conical Tilt revisited

Cryo-Electron Microscopy (cryo-EM) of macromolecular complexes is a fundamental structural biology technique which is expanding at a very fast pace. Key to its success in elucidating the three-dimensional structure of a macromolecular complex, especially of small and non-symmetric ones, is the ability to start from a low resolution map, which is subsequently refined with the actual images collected at the microscope. There are several methods to produce this first structure. Among them, Random Conical Tilt (RCT) plays a prominent role due to its unbiased nature (it can create an initial model based on experimental measurements). In this article, we revise the fundamental mathematical expressions supporting RCT, providing new expressions handling all key geometrical parameters without the need of intermediate operations, leading to improved automation and overall reliability, essential for the success of cryo-EM when analyzing new complexes. We show that the here proposed RCT workflow based on the new formulation performs very well in practical cases, requiring very few image pairs (as low as 13 image pairs in one of our examples) to obtain relevant 3D maps.We thank Dr. Llorca for his support during the acquisition of the C3b images and Dr. Shaikh for his support in the use of Spider for the RCT reconstructions. The authors would like to acknowledge economical support from the Spanish Ministry of Economy and Competitiveness through grants AIC-A-2011-0638 and BIO2013-44647-R, the Comunidad de Madrid through grant CAM (S2010/BMD-2305), as well as a postdoctoral Juan de la Cierva grant with reference JCI-2011-10185 to Javier Vargas. Vahid Abrishami is a holder of La Caixa scholarship and C.O.S. Sorzano is recipient of a Ramon y Cajal fellowship