248 research outputs found
RIBFIND: a web server for identifying rigid bodies in protein structures and to aid flexible fitting into cryo EM maps
No full textMotivation: To better analyze low-resolution cryo electron microscopy maps of macromolecular assemblies, component atomic structures frequently have to be flexibly fitted into them. Reaching an optimal fit and preventing the fitting process from getting trapped in local minima can be significantly improved by identifying appropriate rigid bodies in the fitted component.
Results: Here we present the RIBFIND server, a tool for identifying rigid bodies in protein structures. The server identifies rigid bodies in proteins by calculating spatial proximity between their secondary structural elements.
Availability: The RIBFIND web server and its standalone program are available at http://ribfind.ismb.lon.ac.uk
Flexible fitting of proteins and nucleic acids using Gaussian Mixture Models
No full textFundamental aspect of cryo-electron microscopy (cryo-EM) is deriving atomic models from the 3D reconstructions. A number of advances have been made in detector technology and image processing that has enabled some dramatic improvements in resolution. Still, low resolutions continue to plague the field. Cryo-EM reconstructions, which rely on many images of molecules with different orientations, are limited by the noise and conformational heterogeneity often inherent to the molecules of interest. Classification of these structures into distinct classes, enables insights into the conformational space that they inhabit, and can indeed improve the quality of individual reconstructions. However, even within classes, atomic motions are an unavoidable fact of nature. The inherent noise in low-dose electron imaging, along with the number of particles in the sample, limits to what degree classes can accurately be assigned. It is thus an accepted matter, that many reconstructions will by their very nature contain features from many molecule which are only approximately identical. This manifests itself in heterogeneous resolutions, where fluctuations can be attributed to changes in the local similarity of the averaged molecules.
To build atomic models from these low resolution reconstructions, a common approach known as flexible fitting, employs atomic structures which have been solved using high-resolution techniques. In this thesis, I explore a flexible fitting and refinement method which attempts to improve the interpretability of atomic models by estimating the local resolution of the atoms in the underlying data. Using a Gaussian Mixture Model, a single atomic model is used to describe the experimental map, with atoms modelled as isotropic three-dimensional Gaussians with widths determined from the reconstruction. This allows better interpretation of the atomic structure, as coordinate uncertainty is accounted for. Of course, isomorphic Gaussians are limited in their accuracy at representing atomic motions of bonded atoms. Instead, a second representation is derived by modifying the atomic models in the context of a molecular dynamics simulation with perturbations derived from the local resolution information. This representation, composed of an ensemble of atomic structures was shown to produce an improved fit with the data.
Unfortunately, the initial model which is fitted to the experimental data, often requires significant rearrangement of its coordinates in order to fit the cryo-EM map before local resolutions can be estimated. This is because high resolution starting models are typically derived from X-ray experiments, where crystallisation can result in structural change. Frequently, cryo-EM experiments are elucidating structures which have never been seen before. In such cases, structural predictions are used instead. These can be highly accurate, often down to the level of domains, but may require some rearrangement to better fit the data. Flexible fitting approaches are able to fit such models but require the use of restraints to prevent distortions. In the past, the RIBFIND approach has been successfully used to this end, but has been limited to protein structures. RIBFIND2, which is presented in this thesis is able to decompose RNA structures into rigid bodies which can be restrained during flexible fitting procedures.
Combining the GMM method with these RIBFIND2 restraints enabled a diverse set of structural predictions from the recent CASP15 challenge to be flexibly fit into cryo-EM maps, resulting in models with similar quality to the target structures
College Belonging: How First-Year and First-Generation Students Navigate Campus Life [Review]
No full textFostering a sense of belonging for students has long been considered a crucial component of retention and success for colleges and universities. However, there is no universal definition of what "belonging" actually is. In College Belonging: How First-Year and First-Generation Students Navitage Campus Life, Lisa M. Nunn (2021) delves into what it means for college students "to belong." Through student interviews during their first two years of college, Nunn explores how students define and experience belonging; in doing so, creating a new perspective on what belonging is and how students achieve (or are gifted) belonging.Johnson, C., & Gansemer-Topf, A. (2022). College Belonging: How First-Year and First-Generation Students Navigate Campus Life: [Review]. Journal of College Orientation, Transition, and Retention, 29(2). https://doi.org/10.24926/jcotr.v29i2.4869Copyright (c) 2022 Clayton Johnson, Dr. Ann Gansemer-Topf. This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. Copyright of all articles published in JCOTR belong to the author(s) and are published under a CC BY-NC license. <br
Combined approaches to flexible fitting and assessment in virus capsids undergoing conformational change
No full textFitting of atomic components into electron cryo-microscopy (cryoEM) density maps is routinely used to understand the structure and function of macromolecular machines. Many fitting methods have been developed, but a standard protocol for successful fitting and assessment of fitted models is still to be agreed upon among the experts in the field. Here, we created and tested a protocol that highlights important issues related to homology modelling, density map segmentation, rigid and flexible fitting, as well as the assessment of fits. As part of it, we use two different flexible fitting methods (Flex-EM and iMODfit) and demonstrate how combining the analysis of multiple fits and model assessment could result in an improved model. The protocol is applied to the case of the mature and empty capsids of Coxsackievirus A7 (CAV7) by flexibly fitting homology models into the corresponding cryoEM density maps at 8.2Å and 6.1Å resolution. As a result, and due to the improved homology models (derived from recently solved crystal structures of a close homolog – EV71 capsid – in mature and empty forms), the final models present an improvement over previously published models. In close agreement with the capsid expansion observed in the EV71 structures, the new CAV7 models reveal that the expansion is accompanied by ∼5° counterclockwise rotation of the asymmetric unit, predominantly contributed by the capsid protein VP1. The protocol could be applied not only to viral capsids but also to many other complexes characterised by a combination of atomic structure modelling and cryoEM density fitting
The Power in Groups: Using Cluster Analysis to Critically Quantify Women’s STEM Enrollment
No full textDespite efforts to close the gender gap in science, technology, engineering, and math (STEM), disparities still exist, especially in math intensive STEM (MISTEM) majors. Females and males receive similar academic preparation and overall, perform similarly, yet females continue to enroll in STEM majors less frequently than men. In examining academic preparation, most research considers performance measures individually, ignoring the possible interrelationships between these measures. We address this problem by using hierarchical agglomerative clustering – a statistical technique which allows for identifying groups (i.e., clusters) of students who are similar in multiple factors. We first apply this technique to readily available institutional data to determine if we could identify distinct groups. Results illustrated that it was possible to identify nine unique groups. We then examined differences in STEM enrollment by group and by gender. We found that the proportion of females differed by group, and the gap between males and females also varied by group. Overall, males enrolled in STEM at a higher proportion than females and did so regardless of the strength of their academic preparation. Our results provide a novel yet feasible approach to examining gender differences in STEM enrollment in postsecondary education.This article is published as Gansemer-Topf, Ann et al. "The Power in Groups: Using Cluster Analysis to Critically Quantify Women’s STEM Enrollment" In Advances in Research in STEM Education, edited by Associate Kalogiannakis, Maria Ampartzaki. London: IntechOpen, 2022. 10.5772/intechopen.102881. © 2022 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution 3.0 License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited
Structural investigation of the herpes simplex virus 1 fusion mechanism and the underlaying glycoprotein machinery
No full textA nanobody specific to prefusion glycoprotein B neutralizes HSV-1 and HSV-2
No full textThe nine human herpesviruses, including herpes simplex virus 1 and 2, human cytomegalovirus and Epstein–Barr virus, present a significant burden to global public health1. Their envelopes contain at least ten different glycoproteins, which are necessary for host cell tropism, attachment and entry2. The best conserved among them, glycoprotein B (gB), is essential as it performs membrane fusion by undergoing extensive rearrangements from a prefusion to postfusion conformation. At present, there are no antiviral drugs targeting gB or neutralizing antibodies directed against its prefusion form, because of the difficulty in structurally determining and using this metastable conformation. Here we show the isolation of prefusion-specific nanobodies, one of which exhibits strong neutralizing and cross-species activity. By mutational stabilization we solved the herpes simplex virus 1 gB full-length prefusion structure, which allowed the bound epitope to be determined. Our analyses show the membrane-embedded regions of gB and previously unresolved structural features3,4, including a new fusion loop arrangement, providing insights into the initial conformational changes required for membrane fusion. Binding an epitope spanning three domains, proximal only in the prefusion state, the nanobody keeps wild-type HSV-2 gB in this conformation and enabled its native prefusion structure to be determined. This also indicates the mode of neutralization and an attractive avenue for antiviral interventions
Gefangen im eigenen Ich: Ein psychoanalytischer Vergleich von E.T.A. Hoffmanns \u3cem\u3eDer Sandmann\u3c/em\u3e und \u3cem\u3eDer goldne Topf\u3c/em\u3e
No full textThis thesis is a comparative study of two major works by the German author E.T.A. Hoffmann, Der Sandmann (1818) and Der goldne Topf (1819). Der Sandmann has been analyzed under the filter of psychoanalysis by Freud himself. The goal of this thesis was to analyze whether a psychoanalytical approach can be extended to other works by Hoffmann, showing the same underlying structures even though the content seems to differ widely between the two works at first glance. Der goldne Topf is the text that I chose to compare to Der Sandmann, as both texts tell the story of a student who is caught in a life between reality and fantasy.
Freud\u27s analysis of Der Sandmann is almost completely based on the role of the father in the text. The strongest difficulty in showing the same underlying motivation for the two protagonists, Anselmus and Nathanael, to choose fantasy over reality, death over life, is that there is no apparent father figure in Der goldne Topf. However, by interpreting the two texts on the basis of Freud\u27s psychoanalysis, it can be shown that a father figure is indeed present in both texts, even though it might not seem like it at first.
In chapter 1 of this thesis, I will give an overview over those parts of Freud\u27s theory, which will be of importance in the analyses of the two selected works by Hoffmann, namely narcissism, the oedipus complex and the analysis of dreams. In chapter 2, I interpret Der Sandmann and in chapter 3, Der goldne Topf is analyzed, applying the same theories as far as possible. Finally, in chapter 4, I compare the two works, and I show that many features of the texts can be matched up on the basis of this theory, including the role of the father. So far, Der goldne Topf has never been analyzed exclusively on the basis of Freud\u27s psychoanalysis before and, therefore, the findings of this thesis provide new insights for research on the two texts and on E.T.A. Hoffmann in general
Assessing scoring metrics for AlphaFold2 and AlphaFold3 protein complex predictions
No full textRecent breakthroughs in AI-driven protein structure prediction have revolutionized structural biology, unlocking new possibilities to model complex biomolecular interactions. We evaluated widely used scoring metrics for assessing models predicted by ColabFold with templates, ColabFold without templates, and AlphaFold3. We benchmarked the optimal cutoffs for these assessment scores using a set of 223 heterodimeric, high-resolution protein structures and their predictions. Our results show that ColabFold with templates and AlphaFold3 perform similarly, and both outperform ColabFold without templates. However, the assessment scores perform best on ColabFold without templates. Furthermore, interface-specific scores are more reliable for evaluating protein complex predictions compared to the corresponding global scores. Notably, ipTM and model confidence achieve the best discrimination between correct and incorrect predictions. Based on our results, we developed a weighted combined score, C2Qscore, to improve model quality assessment. We used C2Qscore to analyze dimers from large assemblies solved by cryoEM, revealing potential limitations of the existing metrics when multiple configurations of heterodimers are possible. This study provides insights into the strengths and weaknesses of current scores and offers guidance for improving protein complex model assessment under realistic use case conditions. C2Qscore has been integrated as a tool into our ChimeraX plug-in PICKLUSTER v.2.0 and is also available as a command-line tool on https://gitlab.com/topf-lab/c2qscore
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