ModeHunter: A Package for the Reductionist Analysis, Animation, and Application of Elastic Biomolecular Motion.

ModeHunter is a modular Python software package for the simulation of 3D biophysical motion across spatial resolution scales using modal analysis of elastic networks. It has been curated from our in-house Python scripts over the last 15 years, with a focus on detecting similarities of elastic motion between atomic structures, coarse-grained graphs, and volumetric data obtained from biophysical or biomedical imaging origins, such as electron microscopy or tomography. With ModeHunter, normal modes of biophysical motion can be analyzed with various static visualization techniques or brought to life by dynamics animation in terms of single or multimode trajectories or decoy ensembles.

The combined focal loss and dice loss function improves the segmentation of beta-sheets in medium-resolution cryo-electron-microscopy density maps.

Summary: Although multiple neural networks have been proposed for detecting secondary structures from medium-resolution (5-10 Å) cryo-electron microscopy (cryo-EM) maps, the loss functions used in the existing deep learning networks are primarily based on cross-entropy loss, which is known to be sensitive to class imbalances. We investigated five loss functions: cross-entropy, Focal loss, Dice loss, and two combined loss functions. Using a U-Net architecture in our DeepSSETracer method and a dataset composed of 1355 box-cropped atomic-structure/density-map pairs, we found that a newly designed loss function that combines Focal loss and Dice loss provides the best overall detection accuracy for secondary structures.

Untangling Irregular Actin Cytoskeleton Architectures in Tomograms of the Cell with .

In this work, we established, validated, and optimized a novel computational framework for tracing arbitrarily oriented actin filaments in cryo-electron tomography maps. Our approach was designed for highly complex intracellular architectures in which a long-range cytoskeleton network extends throughout the cell bodies and protrusions. The irregular organization of the actin network, as well as cryo-electron-tomography-specific noise, missing wedge artifacts, and map dimensions call for a specialized implementation that is both robust and efficient.

Spaghetti Tracer: A Framework for Tracing Semiregular Filamentous Densities in 3D Tomograms.

Within cells, cytoskeletal filaments are often arranged into loosely aligned bundles. These fibrous bundles are dense enough to exhibit a certain regularity and mean direction, however, their packing is not sufficient to impose a symmetry between-or specific shape on-individual filaments. This intermediate regularity is computationally difficult to handle because individual filaments have a certain directional freedom, however, the filament densities are not well segmented from each other (especially in the presence of noise, such as in cryo-electron tomography).

Refinement of AlphaFold2 models against experimental and hybrid cryo-EM density maps.

Recent breakthroughs in deep learning-based protein structure prediction show that it is possible to obtain highly accurate models for a wide range of difficult protein targets for which only the amino acid sequence is known. The availability of accurately predicted models from sequences can potentially revolutionise many modelling approaches in structural biology, including the interpretation of cryo-EM density maps. Although atomic structures can be readily solved from cryo-EM maps of better than 4 Å resolution, it is still challenging to determine accurate models from lower-resolution density maps.

Tracing Filaments in Simulated 3D Cryo-Electron Tomography Maps Using a Fast Dynamic Programming Algorithm.

We propose a fast, dynamic programming-based framework for tracing actin filaments in 3D maps of subcellular components in cryo-electron tomography. The approach can identify high-density filament segments in various orientations, but it takes advantage of the arrangement of actin filaments within cells into more or less tightly aligned bundles. Assuming that the tomogram can be rotated such that the filaments can be oriented to be directed in a dominant direction (i.

: Simulation of Filamentous Cryo-Electron Tomograms.

As automated filament tracing algorithms in cryo-electron tomography (cryo-ET) continue to improve, the validation of these approaches has become more incumbent. Having a known ground truth on which to base predictions is crucial to reliably test predicted cytoskeletal filaments because the detailed structure of the filaments in experimental tomograms is obscured by a low resolution, as well as by noise and missing Fourier space wedge artifacts. We present a software tool for the realistic simulation of tomographic maps () based on a known filament trace.

Mechanism for the Unfolding of the TOP7 Protein in Steered Molecular Dynamics Simulations as Revealed by Mutual Information Analysis.

We employed mutual information (MI) analysis to detect motions affecting the mechanical resistance of the human-engineered protein Top7. The results are based on the MI analysis of pair contact correlations measured in steered molecular dynamics (SMD) trajectories and their statistical dependence on global unfolding. This study is the first application of the MI analysis to SMD forced unfolding, and we furnish specific SMD recommendations for the utility of parameters and options in the package.

Cylindrical Similarity Measurement for Helices in Medium-Resolution Cryo-Electron Microscopy Density Maps.

Cryo-electron microscopy (cryo-EM) density maps at medium resolution (5-10 Å) reveal secondary structural features such as α-helices and β-sheets, but they lack the side chain details that would enable a direct structure determination. Among the more than 800 entries in the Electron Microscopy Data Bank (EMDB) of medium-resolution density maps that are associated with atomic models, a wide variety of similarities can be observed between maps and models. To validate such atomic models and to classify structural features, a local similarity criterion, the score, is proposed and evaluated in this study.

Correction of Missing-Wedge Artifacts in Filamentous Tomograms by Template-Based Constrained Deconvolution.

Cryo-electron tomography maps often exhibit considerable noise and anisotropic resolution, due to the low-dose requirements and the missing wedge in Fourier space. These spurious features are visually unappealing and, more importantly, prevent an automated segmentation of geometric shapes, requiring a subjective and labor-intensive manual tracing. We developed a novel computational strategy for objectively denoising and correcting missing-wedge artifacts in homogeneous specimen areas of tomograms, where it is assumed that a template repeats itself across the volume under consideration, as happens in the case of filaments.

A cryo-tomography-based volumetric model of the actin core of mouse vestibular hair cell stereocilia lacking plastin 1.

Electron cryo-tomography allows for high-resolution imaging of stereocilia in their native state. Because their actin filaments have a higher degree of order, we imaged stereocilia from mice lacking the actin crosslinker plastin 1 (PLS1). We found that while stereocilia actin filaments run 13 nm apart in parallel for long distances, there were gaps of significant size that were stochastically distributed throughout the actin core.

Accurate flexible refinement of atomic models against medium-resolution cryo-EM maps using damped dynamics.

Background: Dramatic progress has recently been made in cryo-electron microscopy technologies, which now make possible the reconstruction of a growing number of biomolecular structures to near-atomic resolution. However, the need persists for fitting and refinement approaches that address those cases that require modeling assistance.

Methods: In this paper, we describe algorithms to optimize the performance of such medium-resolution refinement methods.

A Pattern Recognition Tool for Medium-resolution Cryo-EM Density Maps and Low-resolution Cryo-ET Density maps.

Cryo-electron microscopy (Cryo-EM) and cryo-electron tomography (cryo-ET) produce 3-D density maps of biological molecules at a range of resolution levels. Pattern recognition tools are important in distinguishing biological components from volumetric maps with the available resolutions. One of the most distinct characters in density maps at medium (5-10 Å) resolution is the visibility of protein secondary structures.

Tracing Actin Filament Bundles in Three-Dimensional Electron Tomography Density Maps of Hair Cell Stereocilia.

Cryo-electron tomography (cryo-ET) is a powerful method of visualizing the three-dimensional organization of supramolecular complexes, such as the cytoskeleton, in their native cell and tissue contexts. Due to its minimal electron dose and reconstruction artifacts arising from the missing wedge during data collection, cryo-ET typically results in noisy density maps that display anisotropic XY versus Z resolution. Molecular crowding further exacerbates the challenge of automatically detecting supramolecular complexes, such as the actin bundle in hair cell stereocilia.

A Balanced Approach to Adaptive Probability Density Estimation.

Our development of a Fast (Mutual) Information Matching (FIM) of molecular dynamics time series data led us to the general problem of how to accurately estimate the probability density function of a random variable, especially in cases of very uneven samples. Here, we propose a novel Balanced Adaptive Density Estimation (BADE) method that effectively optimizes the amount of smoothing at each point. To do this, BADE relies on an efficient nearest-neighbor search which results in good scaling for large data sizes.

Computing Spatiotemporal Heat Maps of Lipid Electropore Formation: A Statistical Approach.

We extend the multiscale spatiotemporal heat map strategies originally developed for interpreting molecular dynamics simulations of well-structured proteins to liquids such as lipid bilayers and solvents. Our analysis informs the experimental and theoretical investigation of electroporation, that is, the externally imposed breaching of the cell membrane under the influence of an electric field of sufficient magnitude. To understand the nanoscale architecture of electroporation, we transform time domain data of the coarse-grained interaction networks of lipids and solvents into spatial heat maps of the most relevant constituent molecules.

Comparing an Atomic Model or Structure to a Corresponding Cryo-electron Microscopy Image at the Central Axis of a Helix.

Three-dimensional density maps of biological specimens from cryo-electron microscopy (cryo-EM) can be interpreted in the form of atomic models that are modeled into the density, or they can be compared to known atomic structures. When the central axis of a helix is detectable in a cryo-EM density map, it is possible to quantify the agreement between this central axis and a central axis calculated from the atomic model or structure. We propose a novel arc-length association method to compare the two axes reliably.

Comparison of an Atomic Model and Its Cryo-EM Image at the Central Axis of a Helix.

Cryo-electron microscopy (cryo-EM) is an important biophysical technique that produces three-dimensional (3D) density maps at different resolutions. Because more and more models are being produced from cryo-EM density maps, validation of the models is becoming important. We propose a method for measuring local agreement between a model and the density map using the central axis of the helix.

Spatial Heat Maps from Fast Information Matching of Fast and Slow Degrees of Freedom: Application to Molecular Dynamics Simulations.

We introduce a fast information matching (FIM) method for transforming time domain data into spatial images through handshaking between fast and slow degrees of freedom. The analytics takes advantage of the detailed time series available from biomolecular computer simulations, and it yields spatial heat maps that can be visualized on 3D molecular structures or in the form of interaction networks. The speed of our efficient mutual information solver is on the order of a basic Pearson cross-correlation calculation.

Conserved Intramolecular Interactions Maintain Myosin Interacting-Heads Motifs Explaining Tarantula Muscle Super-Relaxed State Structural Basis.

Tarantula striated muscle is an outstanding system for understanding the molecular organization of myosin filaments. Three-dimensional reconstruction based on cryo-electron microscopy images and single-particle image processing revealed that, in a relaxed state, myosin molecules undergo intramolecular head-head interactions, explaining why head activity switches off. The filament model obtained by rigidly docking a chicken smooth muscle myosin structure to the reconstruction was improved by flexibly fitting an atomic model built by mixing structures from different species to a tilt-corrected 2-nm three-dimensional map of frozen-hydrated tarantula thick filament.

Multi-scale Visualization of Molecular Architecture Using Real-Time Ambient Occlusion in Sculptor.

The modeling of large biomolecular assemblies relies on an efficient rendering of their hierarchical architecture across a wide range of spatial level of detail. We describe a paradigm shift currently under way in computer graphics towards the use of more realistic global illumination models, and we apply the so-called ambient occlusion approach to our open-source multi-scale modeling program, Sculptor. While there are many other higher quality global illumination approaches going all the way up to full GPU-accelerated ray tracing, they do not provide size-specificity of the features they shade.

Numerical geometry of map and model assessment.

We are describing best practices and assessment strategies for the atomic interpretation of cryo-electron microscopy (cryo-EM) maps. Multiscale numerical geometry strategies in the Situs package and in secondary structure detection software are currently evolving due to the recent increases in cryo-EM resolution. Criteria that aim to predict the accuracy of fitted atomic models at low (worse than 8Å) and medium (4-8 Å) resolutions remain challenging.

Developing a denoising filter for electron microscopy and tomography data in the cloud.

The low radiation conditions and the predominantly phase-object image formation of cryo-electron microscopy (cryo-EM) result in extremely high noise levels and low contrast in the recorded micrographs. The process of single particle or tomographic 3D reconstruction does not completely eliminate this noise and is even capable of introducing new sources of noise during alignment or when correcting for instrument parameters. The recently developed Digital Paths Supervised Variance (DPSV) denoising filter uses local variance information to control regional noise in a robust and adaptive manner.