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| Open AccessBiomolecular condensate phase diagrams with a combinatorial microdroplet platform
A central concept for characterising phase-separating systems is the phase diagram but generation of such diagrams for biomolecular systems is typically slow and low-throughput. Here the authors describe PhaseScan, a combinatorial droplet microfluidic platform for high-resolution acquisition of multidimensional biomolecular phase diagrams.
- William E. Arter
- , Runzhang Qi
- & Tuomas P. J. Knowles
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Article
| Open AccessCondensates formed by prion-like low-complexity domains have small-world network structures and interfaces defined by expanded conformations
The authors provide a physical description for condensates as complex fluids defined by small-world network structures for their interiors, and of interfaces featuring a preference of locally and globally expanded conformations.
- Mina Farag
- , Samuel R. Cohen
- & Rohit V. Pappu
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Article
| Open AccessVisualizing the transiently populated closed-state of human HSP90 ATP binding domain
To refold client proteins, HSP90 chaperone undergoes large structural rearrangements. Here the authors use NMR and molecular simulation and reveal structure and dynamics of a key functionally relevant metastable state of human HSP90α N-terminal domain.
- Faustine Henot
- , Elisa Rioual
- & Jerome Boisbouvier
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Article
| Open AccessDiscovery and structure of a widespread bacterial ABC transporter specific for ergothioneine
The extent to which human bacterial pathogens broadly utilize ergothioneine as an antioxidant is unknown. Here, authors describe the discovery of a specific transporter for ergothioneine in a respiratory pathogen that is highly conserved among Firmicutes.
- Yifan Zhang
- , Giovanni Gonzalez-Gutierrez
- & David P. Giedroc
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Article
| Open AccessPlanar aggregation of the influenza viral fusion peptide alters membrane structure and hydration, promoting poration
Target membrane poration is a longstanding enigma in virology. Here, authors show that fusion peptides of the influenza virus’ spike protein aggregate, rotate, and tilt in membranes, facilitating peptide-induced poration during viral fusion.
- Amy Rice
- , Sourav Haldar
- & Joshua Zimmerberg
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Article
| Open AccessHydrophobicity of arginine leads to reentrant liquid-liquid phase separation behaviors of arginine-rich proteins
Arginine-rich proteins display reentrant LLPS behavior with changes in salt and temperature due to the hydrophobicity of arginine. Controlling the reentrant LLPS behavior opens avenues for the bioengineering and drug delivery fields.
- Yuri Hong
- , Saeed Najafi
- & Dong Soo Hwang
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Article
| Open AccessA glutamine-based single α-helix scaffold to target globular proteins
Targeting biomedically relevant protein-protein interactions is a long-lasting challenge in medicinal chemistry. Here, the authors develop a single α-helical peptide scaffold that can be tailored to target globular proteins of biomedical interest.
- Albert Escobedo
- , Jonathan Piccirillo
- & Xavier Salvatella
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Article
| Open AccessA simple thermodynamic description of phase separation of Nup98 FG domains
The nuclear pore complex (NPC) barrier is a selective phase assembled from disordered but cohesive FG domains. The authors provide a thermodynamic description of an FG phase that is ultimately simplified and yet closely recapitulates NPC transport selectivity.
- Sheung Chun Ng
- & Dirk Görlich
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Article
| Open AccessQuasAr Odyssey: the origin of fluorescence and its voltage sensitivity in microbial rhodopsins
The authors present an in-depth investigation of excited state dynamics and molecular mechanism of the voltage sensing in microbial rhodopsins. Using a combination of spectroscopic investigations and molecular dynamics simulations, the study proposes the voltage-modulated deprotonation of the chromophore as the key event in the voltage sensing. Thus, molecular constraints that may further improve the fluorescence quantum yield and the voltage sensitivity are presented.
- Arita Silapetere
- , Songhwan Hwang
- & Peter Hegemann
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Article
| Open AccessWater regulates the residence time of Benzamidine in Trypsin
Water is an essential part of any biological system, yet many aspects of its role remain elusive. Here the authors show, in a paradigmatic ligand-protein system, that water modulates the ligand residence time in a complex and non-local way, with possible implications in drug design.
- Narjes Ansari
- , Valerio Rizzi
- & Michele Parrinello
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Article
| Open AccessA blind benchmark of analysis tools to infer kinetic rate constants from single-molecule FRET trajectories
The ability to infer quantitative kinetic information from single-molecule FRET (smFRET) data can be challenging. Here the authors perform a blind benchmark study assessing different analysis tools used to infer kinetic rate constants from smFRET trajectories, testing on simulated and experimental data.
- Markus Götz
- , Anders Barth
- & Sonja Schmid
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Article
| Open AccessStructure and function of H+/K+ pump mutants reveal Na+/K+ pump mechanisms
Here, the authors study ion transport mechanisms by introducing mutations to convert an electroneutral proton/potassium pump into a prototypical electrogenic sodium/potassium pump, explaining their selectivity and phosphorylation mechanisms.
- Victoria C. Young
- , Hanayo Nakanishi
- & Kazuhiro Abe
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Article
| Open AccessMultivalent interactions between molecular components involved in fast endophilin mediated endocytosis drive protein phase separation
Here the authors show that protein phase separation is a key mechanism in cellular receptor internalization via fast endophilin mediated endocytosis (FEME). Phase separation facilitates multivalent FEME-protein assembly in this clathrin-independent pathway.
- Samsuzzoha Mondal
- , Karthik Narayan
- & Tobias Baumgart
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Article
| Open AccessA comprehensive thermodynamic model for RNA binding by the Saccharomyces cerevisiae Pumilio protein PUF4
Traditional genomic methods identify RNA-binding proteins (RBPs) and the genes they regulate, but do not provide predictive models. The authors used an emerging technology to obtain a complete thermodynamic model for RNA binding to the PUF4 RBP.
- Christoph Sadée
- , Lauren D. Hagler
- & Daniel Herschlag
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Article
| Open AccessCross-validation of distance measurements in proteins by PELDOR/DEER and single-molecule FRET
Pulsed electron-electron double resonance spectroscopy (PELDOR/DEER) and single-molecule Förster resonance energy transfer spectroscopy (smFRET) are used to determine conformational changes and probe distances in biological macromolecules. Here the authors compare the methods on a large set of samples.
- Martin F. Peter
- , Christian Gebhardt
- & Gregor Hagelueken
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Article
| Open AccessHeparin induces α-synuclein to form new fibril polymorphs with attenuated neuropathology
The Cryo-EM structures reported in this work reveal how heparin incorporates into α-syn fibril formation to determine fibril polymorphs. This highlights the role of biological polymers in the conformational selection and neuropathological regulation of amyloid fibrils.
- Youqi Tao
- , Yunpeng Sun
- & Dan Li
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Article
| Open AccessNeurotoxic amyloidogenic peptides in the proteome of SARS-COV2: potential implications for neurological symptoms in COVID-19
Here the authors report the formation of toxic clumps of protein, similar to amyloid assemblies found in Alzheimer’s disease and suggest their possible role for some of the neurological symptoms of long-COVID.
- Mirren Charnley
- , Saba Islam
- & Nicholas P. Reynolds
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Article
| Open AccessDynamic arrest and aging of biomolecular condensates are modulated by low-complexity domains, RNA and biochemical activity
Here the authors analyze material properties and aging of active phase-separated condensates by Differential Dynamic Microscopy. Arrested states are promoted by structured RNA. Low-complexity domains and biochemical reaction keep the droplets fluid-like.
- Miriam Linsenmeier
- , Maria Hondele
- & Paolo Arosio
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Article
| Open AccessFluent molecular mixing of Tau isoforms in Alzheimer’s disease neurofibrillary tangles
The tau protein in Alzheimer’s disease contains two isoforms. Using solid-state NMR and seeded growth of isotopically labeled tau, here the authors determined that the two isoforms mix fluently on the molecular level to propagate the AD tau structure.
- Aurelio J. Dregni
- , Pu Duan
- & Mei Hong
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Article
| Open AccessTemplated folding of the RTX domain of the bacterial toxin adenylate cyclase revealed by single molecule force spectroscopy
The authors use optical tweezers to show that the folding of repeats-in-toxin (RTX) block-iv in adenylate cyclase is templated by the folded RTX block-v. The findings suggest a possible mechanism for transmitting the folding signal in the RTX domain.
- Han Wang
- , Guojun Chen
- & Hongbin Li
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Article
| Open AccessStructural anatomy of Protein Kinase C C1 domain interactions with diacylglycerol and other agonists
Protein kinase Cs (PKCs) define a central DAG-sensing node in intracellular phosphoinositide signaling pathways that regulate cell growth, differentiation, apoptosis, and motility. The structures of PKC C1 domain complexes with DAG and 4 agonists reveal the molecular basis of ligand recognition and capture.
- Sachin S. Katti
- , Inna V. Krieger
- & Tatyana I. Igumenova
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Article
| Open AccessCryo-EM structure of human glucose transporter GLUT4
Small solute carriers remain difficult to study by single particle cryo-EM. Here, the authors report the cryo-EM structure of human insulin-responsive glucose transporter GLUT4 (55 kDa) without rigid soluble domains or binders.
- Yafei Yuan
- , Fang Kong
- & Chuangye Yan
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Article
| Open AccessStructural basis of ion – substrate coupling in the Na+-dependent dicarboxylate transporter VcINDY
DASS carboxylate transporters linked to metabolic diseases couple the Na+ gradient to nutrient uptake. Here, the authors unravel the structural basis by which Na+ binding coordinates substrate interactions, revealing a distinct coupling mechanism.
- David B. Sauer
- , Jennifer J. Marden
- & Da-Neng Wang
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Article
| Open AccessFoldamers reveal and validate therapeutic targets associated with toxic α-synuclein self-assembly
Inhibiting alpha-synuclein self-assembly into amyloid structures, associated with Parkinson’s disease, is a potential therapeutic intervention. Here, the authors identify the domains/sequences that are essential for alpha-synuclein aggregation and test the activity of foldamer-based antagonists to identify potential therapeutic targets.
- Jemil Ahmed
- , Tessa C. Fitch
- & Sunil Kumar
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Article
| Open AccessRadiationless mechanism of UV deactivation by cuticle phenolics in plants
Phenolics are abundant in plant cuticles. Here, via transient absorption spectroscopy and quantum chemical calculations, the authors propose a model by which cuticle phenolics provide photoprotection due to ultrafast and non-radiative excited state deactivation combined with fluorescence emission.
- Ana González Moreno
- , Abel de Cózar
- & Antonio Heredia
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Article
| Open AccessMembrane thickness, lipid phase and sterol type are determining factors in the permeability of membranes to small solutes
Membrane permeability of small molecules depends on the composition of the lipid bilayer. Here, authors compare permeability measured on membranes in different physical states and conclude that the yeast membrane exists in a highly ordered phase.
- Jacopo Frallicciardi
- , Josef Melcr
- & Bert Poolman
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Article
| Open AccessA different perspective for nonphotochemical quenching in plant antenna complexes
Nonphotochemical quenching (NPQ) protects photosynthetic complexes from damage due to excess light. Here the authors explore different conformations of the plant CP29 light harvesting complex, showing how protein tuning of carotenoid excitation energies and carotenoid-chlorophyll interactions account for NPQ.
- Edoardo Cignoni
- , Margherita Lapillo
- & Benedetta Mennucci
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Article
| Open AccessRoom-temperature dynamic nuclear polarization enhanced NMR spectroscopy of small biological molecules in water
Dynamic nuclear polarization (DNP) greatly improves the NMR sensitivity, but its implementation in aqueous solutions is challenging. Here the authors demonstrate carbon polarization enhancement via in situ Overhauser DNP in small biomolecules in water at room temperature and high magnetic field.
- Danhua Dai
- , Xianwei Wang
- & Jiafei Mao
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Article
| Open AccessDissection of the amyloid formation pathway in AL amyloidosis
AL amyloidosis is caused by the accumulation of overproduced light chain (LC) fragments as fibrils in patient organs and it is the most prevalent systemic amyloidosis. Here, the authors combine biochemical and biophysical experiments to characterise the lag phase of a patient-derived truncated LC and they identify structural transitions that precede fibril formation.
- Pamina Kazman
- , Ramona M. Absmeier
- & Johannes Buchner
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Article
| Open AccessA previously unrecognized membrane protein in the Rhodobacter sphaeroides LH1-RC photocomplex
Rhodobacter (Rba.) sphaeroides is a model organism for studying bacterial photosynthesis. Here, the authors present the 2.9 Å cryo-EM structure of the monomeric light-harvesting-reaction center core complex from Rba. sphaeroides strain IL106, which revealed the position and conformation of PufX and the presence of an additional component protein-U, an integral membrane protein.
- Kazutoshi Tani
- , Kenji V. P. Nagashima
- & Zheng-Yu Wang-Otomo
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Article
| Open AccessSustained enzymatic activity and flow in crowded protein droplets
Living cells can harvest environmental energy to drive chemical processes. Here the authors design a minimal artificial system that achieves steady states at similar metabolic densities to microorganisms.
- Andrea Testa
- , Mirco Dindo
- & Paola Laurino
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Article
| Open AccessDynamic interactions and Ca2+-binding modulate the holdase-type chaperone activity of S100B preventing tau aggregation and seeding
The calcium binding protein S100B is an abundantly expressed protein in the brain and has neuro-protective functions by inhibiting Aβ aggregation and metal ion toxicity. Here, the authors combine cell biology and biochemical experiments with chemical kinetics and NMR measurements and show that S100B protein is an extracellular Tau chaperone and further characterize the interactions between S100B and Tau.
- Guilherme G. Moreira
- , François-Xavier Cantrelle
- & Cláudio M. Gomes
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Article
| Open AccessStructural basis of terephthalate recognition by solute binding protein TphC
The presence of the gene encoding the solute binding protein TphC has been shown to permit the uptake of terephthalate (TPA), which is the breakdown product of Polyethylene terephthalate (PET) plastic. Here the authors present a structural characterization of TphC in both open and TPA-bound closed conformations.
- Trishnamoni Gautom
- , Dharmendra Dheeman
- & Neil Dixon
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Article
| Open AccessThe Hsc70 disaggregation machinery removes monomer units directly from α-synuclein fibril ends
Molecular chaperones from the Hsp70 family can break up protein aggregates, including amyloids. Here, the authors utilize microfluidic diffusional sizing to assess the mechanism of α-synuclein (αS) disaggregation by the Hsc70–DnaJB1–Apg2 system, and show that single αS molecules are removed directly from the fibril ends.
- Matthias M. Schneider
- , Saurabh Gautam
- & Tuomas P. J. Knowles
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Article
| Open AccessSpontaneous assembly of redox-active iron-sulfur clusters at low concentrations of cysteine
Iron-sulfur (FeS) proteins are involved in electron transfer and CO2 fixation. Here, the authors show that FeS clusters can form spontaneously in the presence of the amino acid cysteine, in conditions similar those expected in Hadean alkaline hydrothermal vents, suggesting a plausible mechanism of their emergence at the origin of life.
- Sean F. Jordan
- , Ioannis Ioannou
- & Nick Lane
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Article
| Open AccessC9orf72-derived arginine-rich poly-dipeptides impede phase modifiers
Nuclear import receptors (NIRs) regulate self-association of RNA-binding proteins as phase modifiers, while C9orf72-derived arginine-rich polydipeptides lead to aberrant phase transitions. Here the authors show in molecular basis how arginine-rich poly-dipeptides impede the ability of NIRs, particularly Kapβ2.
- Hitoki Nanaura
- , Honoka Kawamukai
- & Eiichiro Mori
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Article
| Open AccessSpectroscopic glimpses of the transition state of ATP hydrolysis trapped in a bacterial DnaB helicase
Here, the authors use solid-state NMR and EPR measurements to characterise the ATP hydrolysis transition state of the oligomeric bacterial DnaB helicase from Helicobacter pylori, which was trapped by using aluminium fluoride as a chemical mimic. They identify protein protons that coordinate to the phosphate groups of ADP and DNA and observe that the aluminium fluoride unit is highly mobile and fast-rotating.
- Alexander A. Malär
- , Nino Wili
- & Thomas Wiegand
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Article
| Open AccessDEEP picker is a deep neural network for accurate deconvolution of complex two-dimensional NMR spectra
The analysis of NMR spectra of complex biochemical samples with respect to individual resonances is challenging but critically important. Here, the authors present a deep learning-based method that accelerates this process also for crowded NMR data that are non-trivial to analyze, even by expert NMR spectroscopists.
- Da-Wei Li
- , Alexandar L. Hansen
- & Rafael Brüschweiler
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Article
| Open AccessEndo-lysosomal Aβ concentration and pH trigger formation of Aβ oligomers that potently induce Tau missorting
Aβ oligomers (AβO) are thought to represent the main toxic species in Alzheimer’s disease but very high Aβ concentrations are required to study them in vitro and it remains unknown what role these off-pathway oligomers play in vivo. Here, the authors use a dimeric variant of Aβ termed dimAβ, where two Aβ40 units are linked, which facilitates to study AβO formation kinetics and they observe that Aβ off-pathway oligomer formation is strongly accelerated at endo-lysosomal pH, while amyloid fibril formation is delayed. Furthermore, the authors demonstrate that dimAβ is a disease-relevant model construct for pathogenic AβO formation by showing that dimAβ AβOs target dendritic spines and induce AD-like somatodendritic Tau missorting.
- Marie P. Schützmann
- , Filip Hasecke
- & Wolfgang Hoyer
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Article
| Open AccessNuclear condensates of p300 formed though the structured catalytic core can act as a storage pool of p300 with reduced HAT activity
The histone acetyltransferase p300 mostly localizes to active chromatin; however, some repressed genes marked with H3K27me3 are also bound by p300. Here the authors show p300 is capable of phase separation, which relies on its catalytic core, and that p300 catalytic activity is decreased in phase-separated droplets that co-localize with H3K27me3-marked chromatin.
- Yi Zhang
- , Kyle Brown
- & Tatiana G. Kutateladze
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Article
| Open AccessAllosteric modulation of ghrelin receptor signaling by lipids
The membrane is an integral component of the G protein-coupled receptor signaling machinery. Here authors demonstrate that lipids regulate the signaling efficacy and selectivity of the ghrelin receptor GHSR through specific interactions and bulk effects and observe PIP2 and GM3 induced shifts of the conformational equilibrium of GHSR away from its inactive state.
- Marjorie Damian
- , Maxime Louet
- & Jean-Louis Banères
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Article
| Open AccessA self-organized synthetic morphogenic liposome responds with shape changes to local light cues
The authors generated a Synthetic Morphogenic Membrane System by encapsulating a dynamic microtubule aster and a light-inducible signaling system driven by GTP/ATP chemical potential into cell-sized liposomes. This reconstitution of artificial proto-cells reveals how non-equilibrium phenomena affect cellular information processing in morphogenesis.
- Konstantin Gavriljuk
- , Bruno Scocozza
- & Philippe I. H. Bastiaens
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Article
| Open AccessIce-nucleating proteins are activated by low temperatures to control the structure of interfacial water
Ice-nucleating proteins promote ice formation at high sub-zero temperatures, but the mechanism is still unclear. The authors investigate a model ice-nucleating protein at the air-water interface using vibrational sum frequency generation spectroscopy and simulations, revealing its reorientation at low temperatures, which increases contact with water molecules and promotes their ordering.
- Steven J. Roeters
- , Thaddeus W. Golbek
- & Tobias Weidner
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Article
| Open AccessReentrant liquid condensate phase of proteins is stabilized by hydrophobic and non-ionic interactions
Elucidating the molecular driving forces underlying liquid–liquid phase separation is a key objective for understanding biological function and malfunction. Here the authors show that a wide range of cellular proteins, including FUS, TDP-43, Brd4, Sox2, and Annexin A11, which form condensates at low salt concentrations, can reenter a phase-separated regime at high salt concentrations.
- Georg Krainer
- , Timothy J. Welsh
- & Tuomas P. J. Knowles
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Article
| Open AccessCrystal structure of a photosynthetic LH1-RC in complex with its electron donor HiPIP
The high potential iron-sulfur (HiPIP) proteins are direct electron donors to the light-harvesting-reaction center complexes (LH1-RC) in photosynthetic β- and γ-Proteobacteria. Here, the authors present the 2.9 Å crystal structure of the HiPIP-bound LH1-RC complex from the thermophilic purple sulfur bacterium Thermochromatium tepidum and discuss mechanistic implications for the electron transfer pathway.
- Tomoaki Kawakami
- , Long-Jiang Yu
- & Zheng-Yu Wang-Otomo
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Article
| Open AccessShort-lived metal-centered excited state initiates iron-methionine photodissociation in ferrous cytochrome c
The dissociation mechanism of the heme axial ligand in heme proteins is not yet fully understood. The authors investigate the photodissociation dynamics of the bond between heme Fe and methionine S in ferrous cytochrome c using femtosecond time-resolved X-ray solution scattering and X-ray emission spectroscopy, simultaneously tracking electronic and nuclear structure changes.
- Marco E. Reinhard
- , Michael W. Mara
- & Kelly J. Gaffney
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Article
| Open AccessDesigned folding pathway of modular coiled-coil-based proteins
Coiled-coil protein origami (CCPO) is a strategy for the design of polyhedral cage-shaped protein folds based on coiled-coil (CC) dimer-forming peptides. Here, the authors show that CCPO proteins fold in a multistep process governed by the spatial distance between CC modules in the primary sequence and subsequent folding intermediates, which enables the use of identical CC modules for the CCPO tetrahedron design.
- Jana Aupič
- , Žiga Strmšek
- & Roman Jerala
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Article
| Open AccessStructural basis for a complex I mutation that blocks pathological ROS production
Reactive oxygen species (ROS) production by reverse electron transfer (RET) through complex I is thought to cause tissue damage from heart attacks. Here, the authors combine in vivo work with biochemical and cryo-EM analyses to characterize the effects of a P25L mutation in the ND6 subunit of mitochondrial complex I. They observe that this mutation does not affect oxidative phosphorylation but renders complex I unable to generate ROS by RET: ND6-P25L mice are protected against cardiac ischaemia–reperfusion injury, thus providing evidence for the proposed role of ROS production in myocardial infarction.
- Zhan Yin
- , Nils Burger
- & Judy Hirst
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Article
| Open AccessStructure and dynamics of the drug-bound bacterial transporter EmrE in lipid bilayers
The small proton-coupled transporter EmrE confers multidrug resistance in bacteria. The structure of drug-bound EmrE in phospholipid bilayers is now determined using solid-state NMR. The structure provides detailed insights into the molecular mechanism of substrate recognition by this transporter.
- Alexander A. Shcherbakov
- , Grant Hisao
- & Mei Hong