Crosstalk between regulatory elements in disordered TRPV4 N-terminus modulates lipid-dependent channel activity DOI Creative Commons
Benedikt Goretzki, Christoph Wiedemann, Brett A. McCray

et al.

Nature Communications, Journal Year: 2023, Volume and Issue: 14(1)

Published: July 13, 2023

Abstract Intrinsically disordered regions (IDRs) are essential for membrane receptor regulation but often remain unresolved in structural studies. TRPV4, a member of the TRP vanilloid channel family involved thermo- and osmosensation, has large N-terminal IDR approximately 150 amino acids. With an integrated biology approach, we analyze ensemble TRPV4 network antagonistic regulatory elements it encodes. These modulate activity hierarchical lipid-dependent manner through transient long-range interactions. A highly conserved autoinhibitory patch acts as master regulator by competing with PIP 2 binding to attenuate activity. Molecular dynamics simulations show that loss interaction between -binding site reduces force exerted on structured core TRPV4. This work demonstrates coupled highlights importance IDRs function regulation.

Language: Английский

Versatile and multivalent nanobodies efficiently neutralize SARS-CoV-2 DOI Creative Commons
Yufei Xiang, Sham Nambulli, Zhengyun Xiao

et al.

Science, Journal Year: 2020, Volume and Issue: 370(6523), P. 1479 - 1484

Published: Nov. 6, 2020

Nanobodies that neutralize Monoclonal antibodies bind to the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) show therapeutic promise but must be produced in mammalian cells and need delivered intravenously. By contrast, single-domain called nanobodies can bacteria or yeast, their stability may enable aerosol delivery. Two papers now report tightly efficiently SARS-CoV-2 cells. Schoof et al. screened a yeast surface display synthetic Xiang anti-spike by llama. Both groups identified highly potent lock an inactive conformation. Multivalent constructs selected achieved even more neutralization. Science , this issue p. 1473 1479

Language: Английский

Citations

406
Cross-Linking Mass Spectrometry for Investigating Protein Conformations and Protein–Protein Interactions─A Method for All Seasons DOI Creative Commons
Lolita Piersimoni, Panagiotis L. Kastritis,

Christian Arlt

et al.

Chemical Reviews, Journal Year: 2021, Volume and Issue: 122(8), P. 7500 - 7531

Published: Nov. 19, 2021

Mass spectrometry (MS) has become one of the key technologies structural biology. In this review, contributions chemical cross-linking combined with mass (XL-MS) for studying three-dimensional structures proteins and investigating protein–protein interactions are outlined. We summarize most important reagents, software tools, XL-MS workflows highlight prominent examples characterizing proteins, their assemblies, interaction networks in vitro vivo. Computational modeling plays a crucial role deriving 3D-structural information from data. Integrating other techniques biology, such as cryo-electron microscopy, been successful addressing biological questions that to date could not be answered. is therefore expected play an increasingly biology future.

Language: Английский

Citations

193
Comprehensive structure and functional adaptations of the yeast nuclear pore complex DOI Creative Commons
Christopher W. Akey, Digvijay Singh, Christna Ouch

et al.

Cell, Journal Year: 2022, Volume and Issue: 185(2), P. 361 - 378.e25

Published: Jan. 1, 2022

Nuclear pore complexes (NPCs) mediate the nucleocytoplasmic transport of macromolecules. Here we provide a structure isolated yeast NPC in which inner ring is resolved by cryo-EM at sub-nanometer resolution to show how flexible connectors tie together different structural and functional layers. These may be targets for phosphorylation regulated disassembly cells with an open mitosis. Moreover, some nucleoporin pairs factors have similar interaction motifs, suggests evolutionary mechanistic link between assembly transport. We evidence three major variants that foreshadow specializations nuclear periphery. Cryo-electron tomography extended these studies, providing model situ radially expanded ring. Our comprehensive reveals features basket central transporter, role lumenal Pom152 restricting dilation, highlights plasticity required

Language: Английский

Citations

144
Better, Faster, Cheaper: Recent Advances in Cryo–Electron Microscopy DOI Creative Commons
Eugene Chua, Joshua H. Mendez, Micah Rapp

et al.

Annual Review of Biochemistry, Journal Year: 2022, Volume and Issue: 91(1), P. 1 - 32

Published: March 23, 2022

Cryo-electron microscopy (cryo-EM) continues its remarkable growth as a method for visualizing biological objects, which has been driven by advances across the entire pipeline. Developments in both single-particle analysis and situ tomography have enabled more structures to be imaged determined better resolutions, at faster speeds, with scientists having improved access. This review highlights recent each stageof cryo-EM pipeline provides examples of how these techniques used investigate real-world problems, including antibody development against SARS-CoV-2 spike during COVID-19 pandemic.

Language: Английский

Citations

105
Reliability and accuracy of single-molecule FRET studies for characterization of structural dynamics and distances in proteins DOI Creative Commons
Ganesh Agam, Christian Gebhardt, Milana Popara

et al.

Nature Methods, Journal Year: 2023, Volume and Issue: 20(4), P. 523 - 535

Published: March 27, 2023

Abstract Single-molecule Förster-resonance energy transfer (smFRET) experiments allow the study of biomolecular structure and dynamics in vitro vivo. We performed an international blind involving 19 laboratories to assess uncertainty FRET for proteins with respect measured efficiency histograms, determination distances, detection quantification structural dynamics. Using two protein systems distinct conformational changes dynamics, we obtained ≤0.06, corresponding interdye distance precision ≤2 Å accuracy ≤5 Å. further discuss limits detecting fluctuations this range how identify dye perturbations. Our work demonstrates ability smFRET simultaneously measure distances avoid averaging realistic systems, highlighting its importance expanding toolbox integrative biology.

Language: Английский

Citations

82
The HADDOCK2.4 web server for integrative modeling of biomolecular complexes DOI
Rodrigo V. Honorato, Mikaël Trellet, Brian Jiménez‐García

et al.

Nature Protocols, Journal Year: 2024, Volume and Issue: unknown

Published: June 17, 2024

Language: Английский

Citations

79
A quantitative map of nuclear pore assembly reveals two distinct mechanisms DOI Creative Commons
Shotaro Otsuka, Jeremy O. B. Tempkin, Wanlu Zhang

et al.

Nature, Journal Year: 2023, Volume and Issue: 613(7944), P. 575 - 581

Published: Jan. 4, 2023

Understanding how the nuclear pore complex (NPC) is assembled of fundamental importance to grasp mechanisms behind its essential function and understand role during evolution eukaryotes

Language: Английский

Citations

46
Integrating cellular electron microscopy with multimodal data to explore biology across space and time DOI Creative Commons
Caitlyn L McCafferty, Sven Klumpe, Rommie E. Amaro

et al.

Cell, Journal Year: 2024, Volume and Issue: 187(3), P. 563 - 584

Published: Feb. 1, 2024

Biology spans a continuum of length and time scales. Individual experimental methods only glimpse discrete pieces this spectrum but can be combined to construct more holistic view. In Review, we detail the latest advancements in volume electron microscopy (vEM) cryo-electron tomography (cryo-ET), which together visualize biological complexity across scales from organization cells large tissues molecular details inside native cellular environments. addition, discuss emerging methodologies for integrating three-dimensional (3DEM) imaging with multimodal data, including fluorescence microscopy, mass spectrometry, single-particle analysis, AI-based structure prediction. This multifaceted approach fills gaps continuum, providing functional context, spatial organization, identity, interactions. We conclude perspective on incorporating diverse data into computational simulations that further bridge extend while dimension time.

Language: Английский

Citations

41
CombFold: predicting structures of large protein assemblies using a combinatorial assembly algorithm and AlphaFold2 DOI Creative Commons
Ben Shor, Dina Schneidman‐Duhovny

Nature Methods, Journal Year: 2024, Volume and Issue: 21(3), P. 477 - 487

Published: Feb. 7, 2024

Abstract Deep learning models, such as AlphaFold2 and RosettaFold, enable high-accuracy protein structure prediction. However, large complexes are still challenging to predict due their size the complexity of interactions between multiple subunits. Here we present CombFold, a combinatorial hierarchical assembly algorithm for predicting structures utilizing pairwise subunits predicted by AlphaFold2. CombFold accurately (TM-score >0.7) 72% among top-10 predictions in two datasets 60 large, asymmetric assemblies. Moreover, structural coverage was 20% higher compared corresponding Protein Data Bank entries. We applied method on from Complex Portal with known stoichiometry but without obtained high-confidence predictions. supports integration distance restraints based crosslinking mass spectrometry fast enumeration possible complex stoichiometries. CombFold’s high accuracy makes it promising tool expanding beyond monomeric proteins.

Language: Английский

Citations

39
The molecular architecture of the nuclear basket DOI Creative Commons
Digvijay Singh, Neelesh Soni, Joshua Hutchings

et al.

Cell, Journal Year: 2024, Volume and Issue: 187(19), P. 5267 - 5281.e13

Published: Aug. 9, 2024

The nuclear pore complex (NPC) is the sole mediator of nucleocytoplasmic transport. Despite great advances in understanding its conserved core architecture, peripheral regions can exhibit considerable variation within and between species. One such structure cage-like basket. crucial roles mRNA surveillance chromatin organization, an architectural has remained elusive. Using in-cell cryo-electron tomography subtomogram analysis, we explored NPC's structural variations basket across fungi (yeast; S. cerevisiae), mammals (mouse; M. musculus), protozoa (T. gondii). integrative modeling, computed a model yeast that revealed how hub nucleoporins (Nups) ring binds to basket-forming Mlp/Tpr proteins: coiled-coil domains form struts basket, while their unstructured termini constitute distal densities, which potentially serve as docking site for preprocessing before

Language: Английский

Citations

23