Electron Microscopy and Single Molecule Studies of Biomolecular Structure and Dynamics

A special issue of Micromachines (ISSN 2072-666X). This special issue belongs to the section "B:Biology and Biomedicine".

Deadline for manuscript submissions: 30 May 2024 | Viewed by 9509

Special Issue Editor


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Guest Editor
1. Institute for Quantitative Biomedicine, Department of Biochemistry & Microbiology, Rutgers University, Piscataway, NJ, USA
2. CryoEMcorp, Bridgewater, NJ, USA
Interests: biophysics; structural biology; cryo-electron microscopy (cryo-EM); correlative light and electron microscopy (CLEM); single-molecule microscopy; DNA replication & repair; mitochondrial disorders; cancer

Special Issue Information

Dear Colleagues,

Technical innovation of the last decade has transformed the field of biophysics by bringing remarkable insight into the mechanisms employed by bio-micromachines to sustain life. Several novel techniques have been developed that allow studies of biomolecular structure and dynamics at unpreceded spatial and temporal resolution. Cryo-electron microscopy (cryo-EM) has revolutionized structural biology. With recent advances in direct electron detectors, improvements in microscope optics, and developments of novel computational algorithms employing artificial intelligence (AI), it is now possible to obtain so-far unattainable atomic biomolecular structures of paramount importance for basic science and the pharmaceutical industry. Advances in single-molecule biophysics allow for the manipulation and study of individual biomolecules without the necessity for ensemble averaging, shedding light on transient intermediate states and molecular fluctuations.

Finally, novel methods such as correlative light and electron microscopy (CLEM), time-resolved cryo-EM, and liquid phase EM (LPEM) have emerged, enabling the correlation of molecular dynamics with structure. This Special Issue entitled “Electron Microscopy and Single Molecule Studies of Biomolecular Structure and Dynamics” seeks to showcase research papers, communications, and review articles that focus on the effective integration and application of recently developed biophysical tools.

Dr. Arkadiusz Kulczyk
Guest Editor

Manuscript Submission Information

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Keywords

  • cryo-electron microscopy (cryo-EM)
  • cryo-electron tomography (cryo-ET)
  • cryo-focused ion beam scanning electron microscopy (cryo-FIB SEM)
  • liquid phase electron microscopy (LPEM)
  • microcrystal electron diffraction (MicroED)
  • novel algorithms, including artificial intelligence, to study protein structure and dynamics
  • optical tweezers
  • single-molecule studies of physical properties of biomolecules
  • super-resolution microscopy
  • time-resolved cryo-electron microscopy

Published Papers (4 papers)

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Research

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17 pages, 6863 KiB  
Article
Delineating Conformational Variability in Small Protein Structures Using Combinatorial Refinement Strategies
by Deborah F. Kelly, G M Jonaid, Liam Kaylor, Maria J. Solares, Samantha Berry, Liza-Anastasia DiCecco, William Dearnaley and Michael Casasanta
Micromachines 2023, 14(10), 1869; https://doi.org/10.3390/mi14101869 - 29 Sep 2023
Cited by 1 | Viewed by 2641
Abstract
As small protein assemblies and even small proteins are becoming more amenable to cryo-Electron Microscopy (EM) structural studies, it is important to consider the complementary dynamic information present in the data. Current computational strategies are limited in their ability to resolve minute differences [...] Read more.
As small protein assemblies and even small proteins are becoming more amenable to cryo-Electron Microscopy (EM) structural studies, it is important to consider the complementary dynamic information present in the data. Current computational strategies are limited in their ability to resolve minute differences among low molecular weight entities. Here, we demonstrate a new combinatorial approach to delineate flexible conformations among small proteins using real-space refinement applications. We performed a meta-analysis of structural data for the SARS CoV-2 Nucleocapsid (N) protein using a combination of rigid-body refinement and simulated annealing methods. For the N protein monomer, we determined three new flexible conformers with good stereochemistry and quantitative comparisons provided new evidence of their dynamic properties. A similar analysis performed for the N protein dimer showed only minor structural differences among the flexible models. These results suggested a more stable view of the N protein dimer than the monomer structure. Taken together, the new computational strategies can delineate conformational changes in low molecular weight proteins that may go unnoticed by conventional assessments. The results also suggest that small proteins may be further stabilized for structural studies through the use of solution components that limit the movement of external flexible regions. Full article
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24 pages, 3249 KiB  
Article
Performance and Quality Comparison of Movie Alignment Software for Cryogenic Electron Microscopy
by David Střelák, Daniel Marchán, José María Carazo and Carlos O. S. Sorzano
Micromachines 2023, 14(10), 1835; https://doi.org/10.3390/mi14101835 - 26 Sep 2023
Viewed by 1342
Abstract
Cryogenic electron microscopy (Cryo-EM) has been established as one of the key players in structural biology. It can reconstruct a 3D model of a sample at a near-atomic resolution. With the increasing number of facilities, faster microscopes, and new imaging techniques, there is [...] Read more.
Cryogenic electron microscopy (Cryo-EM) has been established as one of the key players in structural biology. It can reconstruct a 3D model of a sample at a near-atomic resolution. With the increasing number of facilities, faster microscopes, and new imaging techniques, there is a growing demand for algorithms and programs able to process the so-called movie data produced by the microscopes in real time while preserving a high resolution and maximal information. In this article, we conduct a comparative analysis of the quality and performance of the most commonly used software for movie alignment. More precisely, we compare the most recent versions of FlexAlign (Xmipp v3.23.03), MotionCor2 (v1.6.4), Relion MotionCor (v4.0-beta), Warp (v1.0.9), and CryoSPARC (v4.0.3). We tested the quality of the alignment using generated phantom data, as well as real datasets, comparing the alignment precision, power spectra density, and performance scaling of each program. Full article
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Review

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26 pages, 3442 KiB  
Review
Novel Artificial Intelligence-Based Approaches for Ab Initio Structure Determination and Atomic Model Building for Cryo-Electron Microscopy
by Megan C. DiIorio and Arkadiusz W. Kulczyk
Micromachines 2023, 14(9), 1674; https://doi.org/10.3390/mi14091674 - 27 Aug 2023
Cited by 2 | Viewed by 2005
Abstract
Single particle cryo-electron microscopy (cryo-EM) has emerged as the prevailing method for near-atomic structure determination, shedding light on the important molecular mechanisms of biological macromolecules. However, the inherent dynamics and structural variability of biological complexes coupled with the large number of experimental images [...] Read more.
Single particle cryo-electron microscopy (cryo-EM) has emerged as the prevailing method for near-atomic structure determination, shedding light on the important molecular mechanisms of biological macromolecules. However, the inherent dynamics and structural variability of biological complexes coupled with the large number of experimental images generated by a cryo-EM experiment make data processing nontrivial. In particular, ab initio reconstruction and atomic model building remain major bottlenecks that demand substantial computational resources and manual intervention. Approaches utilizing recent innovations in artificial intelligence (AI) technology, particularly deep learning, have the potential to overcome the limitations that cannot be adequately addressed by traditional image processing approaches. Here, we review newly proposed AI-based methods for ab initio volume generation, heterogeneous 3D reconstruction, and atomic model building. We highlight the advancements made by the implementation of AI methods, as well as discuss remaining limitations and areas for future development. Full article
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24 pages, 3136 KiB  
Review
Exploring the Structural Variability of Dynamic Biological Complexes by Single-Particle Cryo-Electron Microscopy
by Megan C. DiIorio and Arkadiusz W. Kulczyk
Micromachines 2023, 14(1), 118; https://doi.org/10.3390/mi14010118 - 31 Dec 2022
Cited by 5 | Viewed by 2513
Abstract
Biological macromolecules and assemblies precisely rearrange their atomic 3D structures to execute cellular functions. Understanding the mechanisms by which these molecular machines operate requires insight into the ensemble of structural states they occupy during the functional cycle. Single-particle cryo-electron microscopy (cryo-EM) has become [...] Read more.
Biological macromolecules and assemblies precisely rearrange their atomic 3D structures to execute cellular functions. Understanding the mechanisms by which these molecular machines operate requires insight into the ensemble of structural states they occupy during the functional cycle. Single-particle cryo-electron microscopy (cryo-EM) has become the preferred method to provide near-atomic resolution, structural information about dynamic biological macromolecules elusive to other structure determination methods. Recent advances in cryo-EM methodology have allowed structural biologists not only to probe the structural intermediates of biochemical reactions, but also to resolve different compositional and conformational states present within the same dataset. This article reviews newly developed sample preparation and single-particle analysis (SPA) techniques for high-resolution structure determination of intrinsically dynamic and heterogeneous samples, shedding light upon the intricate mechanisms employed by molecular machines and helping to guide drug discovery efforts. Full article
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