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| Open AccessRING domains act as both substrate and enzyme in a catalytic arrangement to drive self-anchored ubiquitination
The mechanism by which RING E3-anchored ubiquitin chains are formed is not well understood. Here, the authors solve a crystal structure of the RING E3 enzyme TRIM21 trapped in the process of self-anchored chain elongation and provide biochemical and cellular insights into the mechanism of ubiquitin conjugation.
- Leo Kiss
- , Dean Clift
- & Leo C. James
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Article
| Open AccessKRAS interaction with RAF1 RAS-binding domain and cysteine-rich domain provides insights into RAS-mediated RAF activation
The molecular details of the RAS-RAF interaction are still not fully understood. Here, the authors present crystal structures of wild-type and mutant KRAS in complex with the RAS-binding and membrane-interacting cysteine-rich domains of RAF1, and propose a model of the membrane-bound RAS-RAF complex.
- Timothy H. Tran
- , Albert H. Chan
- & Dhirendra K. Simanshu
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Article
| Open AccessTranscription activation by a sliding clamp
Transcription activation of late genes in T4 bacteriophage requires the promoter specificity factor gp55, the coactivator gp33 and the sliding clamp gp45. Here, the authors provide structural insights into gp45- dependent transcription activation by determining the cryo-EM structures of a gp55-dependent RNA polymerase (RNAP)-promoter open complex and of an intact gp45-dependent transcription activation complex.
- Jing Shi
- , Aijia Wen
- & Yu Feng
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Article
| Open AccessEltrombopag directly inhibits BAX and prevents cell death
The BCL-2 family protein BAX functions to regulate mitochondria-driven cell death. Here the authors show that the drug Eltrombopag inhibits BAX and prevents apoptosis induced by cytotoxic stimuli.
- Adam Z. Spitz
- , Emmanouil Zacharioudakis
- & Evripidis Gavathiotis
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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 AccessStructure of photosystem I-LHCI-LHCII from the green alga Chlamydomonas reinhardtii in State 2
Photosystems (PS) I and II undergo state transitions to optimize photosynthesis and photoprotection. Here the authors report a cryo-electron microscopy structure of the state 2 PSI-LHCI-LHCII supercomplex from C. reinhardtii revealing subunit organization and possible pathways of energy transfer.
- Zihui Huang
- , Liangliang Shen
- & Guangye Han
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Article
| Open AccessAsymmetric opening of the homopentameric 5-HT3A serotonin receptor in lipid bilayers
Pentameric ligand-gated ion channels (pLGICs) are key players in neurotransmission and have been shown to be modulated by the lipid environment, however the underlying mechanism is not well understood. Here, the authors report structures of the pLGIC 5-HT3A serotonin receptor reconstituted into lipid bilayer discs and reveal lipid–protein interactions as well as asymmetric activation of the homopentameric receptor.
- Yingyi Zhang
- , Patricia M. Dijkman
- & Mikhail Kudryashev
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Article
| Open AccessStructural and biophysical correlation of anti-NANP antibodies with in vivo protection against P. falciparum
The most advanced P. falciparum circumsporozoite protein (PfCSP)-based malaria vaccine confers partial protection. Here, Pholcharee et al. present crystal structures, binding affinities/kinetics, and in vivo protection of 8 anti-NANP antibodies to understand in vivo protection of PfCSP-targeting antibodies.
- Tossapol Pholcharee
- , David Oyen
- & Ian A. Wilson
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Article
| Open AccessHPF1 remodels the active site of PARP1 to enable the serine ADP-ribosylation of histones
Once DNA breaks occur, poly(ADP-ribose) polymerase 1 (PARP1) ADP-ribosylates itself and other DNA repair factors to initiate the repair process. Here, the authors resolve the crystal structures of mouse and human HPF1, and human HPF1/PARP1 complex proving insights into PARP1 regulation.
- Fa-Hui Sun
- , Peng Zhao
- & Cai-Hong Yun
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Article
| Open AccessAA amyloid fibrils from diseased tissue are structurally different from in vitro formed SAA fibrils
Systemic AA amyloidosis is a protein misfolding disease caused by the formation of amyloid fibrils from serum amyloid A (SAA) protein. Here, the authors present the cryo-EM structures of AA amyloid fibrils isolated from mouse tissue and in vitro formed fibrils, which differ in their structures and they also show that the ex vivo fibrils are more resistant to proteolysis than the in vitro fibrils and propose that pathogenic amyloid fibrils might originate from proteolytic selection.
- Akanksha Bansal
- , Matthias Schmidt
- & Marcus Fändrich
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Article
| Open AccessRegulatory inter-domain interactions influence Hsp70 recruitment to the DnaJB8 chaperone
The Hsp70/Hsp40 system plays an important role in maintaining cellular proteostasis but so far it is not well understood how Hsp70 proteins are recruited to specific Hsp40 co-chaperones. Here, the authors combine biochemical and biophysical approaches to characterise the oligomeric mammalian Hsp40 DnaJB8. They identify an intra-oligomer DnaJB8 interaction between the N-terminal J-Domain and the C-terminal domain that occludes the J-Domain surface that binds Hsp70 and propose a model for DnaJB8-Hsp70 recruitment.
- Bryan D. Ryder
- , Irina Matlahov
- & Lukasz A. Joachimiak
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Article
| Open AccessMechanisms of feedback inhibition and sequential firing of active sites in plant aspartate transcarbamoylase
Aspartate transcarbamoylase acts in de novo pyrimidine biosynthesis and in plants is regulated by feedback inhibition via uridine 5-monophosphate (UMP). Here Bellin et al. describe the structural basis for this feedback inhibition, showing that UMP blocks the active site by binding to a plant specific UMP recognition loop.
- Leo Bellin
- , Francisco Del Caño-Ochoa
- & Santiago Ramón-Maiques
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Article
| Open AccessSelf-assembly and regulation of protein cages from pre-organised coiled-coil modules
Coiled-coil protein origami is a strategy for the de novo design of polypeptide nanostructures based on coiled-coil dimer forming peptides, where a single chain protein folds into a polyhedral cage. Here, the authors design a single-chain triangular bipyramid and also demonstrate that the bipyramid can be self-assembled as a heterodimeric complex, comprising pre-defined subunits.
- Fabio Lapenta
- , Jana Aupič
- & Roman Jerala
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Article
| Open AccessStructural mechanism of bivalent histone H3K4me3K9me3 recognition by the Spindlin1/C11orf84 complex in rRNA transcription activation
Spindlin1 is an epigenetic reader that facilitates ribosomal RNA transcription. Here the authors reveal in vitro and structural evidence suggesting that Spindlin1 acts together with C11orf84 to recognize noncanonical bivalent mark of trimethylated lysine 4 and lysine 9 present on histone H3 tail (H3K4me3K9me3).
- Yongming Du
- , Yinxia Yan
- & Chengmin Qian
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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 AccessFunctional and structural characterization of a flavoprotein monooxygenase essential for biogenesis of tryptophylquinone cofactor
An important type of post-translational protein modification is the conversion of peptidyl amino acid into enzyme cofactor. Here, the authors report functional and structural characterization of a flavoprotein monooxygenase essential for biosynthesis of cysteine tryptophylquinone (CTQ) cofactor.
- Toshinori Oozeki
- , Tadashi Nakai
- & Toshihide Okajima
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Article
| Open AccessThe docking of synaptic vesicles on the presynaptic membrane induced by α-synuclein is modulated by lipid composition
α-Synuclein is a presynaptic protein whose aberrant aggregation is associated with Parkinson’s disease. Here, the authors show how αSynuclein-induced docking of synaptic vesicles is modulated by the lipid composition changes typically observed in neurodegeneration using an in vitro system.
- Wing K. Man
- , Bogachan Tahirbegi
- & Giuliana Fusco
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Article
| Open AccessCryo-EM structures of engineered active bc1-cbb3 type CIII2CIV super-complexes and electronic communication between the complexes
Respiratory chains generate the proton motive force used for ATP synthesis. Cryo-EM structures of functional respiratory CIII2CIV supercomplex and native CIII2 from Rhodobacter capsulatus provide insight into CIII2CIV assembly and respiratory electron transport pathways in Gram-negative bacteria.
- Stefan Steimle
- , Trevor van Eeuwen
- & Fevzi Daldal
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Article
| Open AccessCryo-EM reveals structural breaks in a patient-derived amyloid fibril from systemic AL amyloidosis
Systemic AL amyloidosis is a protein misfolding disease caused by the aggregation and fibrillation of immunoglobulin light chains (LCs). Here, the authors present the cryo-EM structures of λ3 LC-derived amyloid fibrils that were isolated from patient tissue and they observe structural breaks, where the two different fibril structures co-exist at different z-axial positions within the same fibril.
- Lynn Radamaker
- , Julian Baur
- & Marcus Fändrich
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Article
| Open AccessStructural basis for the biosynthesis of lovastatin
Biosynthesis of the statin precursor lovastatin depends on the LovB–LovC megasynthase complex. Here, the authors present cryoEM structures of LovB–LovC and core LovB, providing structural insights into the catalytic cycle underlying lovastatin production.
- Jialiang Wang
- , Jingdan Liang
- & Zhijun Wang
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Article
| Open AccessCross-linking mass spectrometry uncovers protein interactions and functional assemblies in synaptic vesicle membranes
Synaptic vesicles store neurotransmitters and fuse with the pre-synaptic membrane when an action potential arrives at the nerve terminal. Here authors apply cross-linking mass spectrometry to study interactions of synaptic vesicle proteins and describe a protein network of vesicle sub-populations and functional assemblies.
- Sabine Wittig
- , Marcelo Ganzella
- & Carla Schmidt
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Article
| Open AccessIncorporation of sensing modalities into de novo designed fluorescence-activating proteins
Fluorescent protein reporters based on GFP exist, but have intrinsic disadvantages. Here the authors incorporate pH, Ca2+ and protein–protein interaction sensing modalities into de novo designed mini-fluorescence-activating proteins (mFAPs), with increased photostability and smaller size, which bind a range of DFHBI chromophore variants.
- Jason C. Klima
- , Lindsey A. Doyle
- & David Baker
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Article
| Open AccessStructure and binding properties of Pangolin-CoV spike glycoprotein inform the evolution of SARS-CoV-2
It has been suggested that pangolin coronaviruses may be the origin of SARS-CoV-2. Here the authors show that the Pangolin-CoV spike is structurally closely related to the closed form of SARS-CoV-2 spike and exhibits similar binding properties to human and pangolin ACE2; although neither spike binds bat ACE2.
- Antoni G. Wrobel
- , Donald J. Benton
- & Steven J. Gamblin
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Article
| Open AccessStructures of active-state orexin receptor 2 rationalize peptide and small-molecule agonist recognition and receptor activation
Agonists of the orexin receptor 2 (OX2R) show promise in the treatment of narcolepsy. Cryo-EM structures of active-state OX2R bound to an endogenous peptide agonist and a small-molecule agonist suggest a molecular mechanism that rationalizes both receptor activation and inhibition.
- Chuan Hong
- , Noel J. Byrne
- & Kaspar Hollenstein
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Article
| Open AccessPathological conformations of disease mutant Ryanodine Receptors revealed by cryo-EM
Ryanodine Receptors (RyRs) release Ca2+ from the endoplasmic and sarcoplasmic reticulum. Mutations in RyR are linked to malignant hyperthermia (MH), myopathies, and arrhythmias. Here, a collection of cryoEM structures provides insights into the molecular consequences of MHrelated RyR mutation R615C, and how apoCaM opens RyR1.
- Kellie A. Woll
- , Omid Haji-Ghassemi
- & Filip Van Petegem
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Article
| Open AccessCryo-EM structural analysis of FADD:Caspase-8 complexes defines the catalytic dimer architecture for co-ordinated control of cell fate
The core FADD:Caspase-8 complex and its regulatory partners, such as the cell death inhibitor c-FLIP, coordinate cell fate. Here authors present the structure of full-length procaspase-8 in a complex with FADD and reveal how recruitment of c-FLIPS into this complex inhibits Caspase-8 activity.
- Joanna L. Fox
- , Michelle A. Hughes
- & Marion MacFarlane
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Article
| Open AccessStructural elements in the flexible tail of the co-chaperone p23 coordinate client binding and progression of the Hsp90 chaperone cycle
p23 is a co-chaperone of Hsp90 but its mode of action is mechanistically not well understood. Here, the authors combine in vitro and yeast in vivo assays, biochemical measurements and NMR experiments to characterize p23 and identify two conserved helical elements in the intrinsically disordered C-terminal tail of p23 that together with the folded domain of p23 regulate the Hsp90 ATPase activity and affect the binding and maturation of Hsp90 clients.
- Maximilian M. Biebl
- , Abraham Lopez
- & Johannes Buchner
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Article
| Open AccessThe mechanism of the nucleo-sugar selection by multi-subunit RNA polymerases
RNA and DNA polymerases need to discriminate efficiently against closely related nucleotide triphosphate substrates. Here, the authors show that a conserved Arg residue is the major determinant of selectivity against deoxyribonucleoside substrates by multisubunit RNA polymerases.
- Janne J. Mäkinen
- , Yeonoh Shin
- & Georgiy A. Belogurov
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Article
| Open AccessGating the pore of the calcium-activated chloride channel TMEM16A
The binding of cytoplasmic Ca2+ to the anion-selective channel TMEM16A triggers a conformational change around its binding site that is coupled to the release of a gate at the constricted neck. Here authors use cryo-EM and electrophysiology to identify three hydrophobic residues at the intracellular entrance of the neck as constituents of this gate.
- Andy K. M. Lam
- , Jan Rheinberger
- & Raimund Dutzler
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Article
| Open AccessDiscovery of an ene-reductase for initiating flavone and flavonol catabolism in gut bacteria
Flavonoids are abundant polyphenols in plants but it is not well understood how their metabolism is initiated by microbes in the human gut. Here, the authors identify and characterise an ene-reductase from the gut bacterium, Flavonifractor plautii ATCC 49531 that catalyses the hydrogenation of the C2–C3 double bond of flavones and flavonols and present its crystal structure.
- Gaohua Yang
- , Sen Hong
- & Yang Gu
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Article
| Open AccessRecognition of non-CpG repeats in Alu and ribosomal RNAs by the Z-RNA binding domain of ADAR1 induces A-Z junctions
ADAR1 is an interferon-induced enzyme that catalyzes editing of adenine to inosine across the transcriptome as part of the immune response. Here the authors establish how ADAR1 recognizes non-CpG RNA sequences to facilitate the formation of A-Z junctions.
- Parker J. Nichols
- , Shaun Bevers
- & Beat Vögeli
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Article
| Open AccessConserved strategies of RNA polymerase I hibernation and activation
RNA polymerase (Pol) I transcribes the ribosomal RNA precursor in eukaryotes. Here, the authors present three cryo-EM structure of S. pombe Pol I in different functional states among them a dimer structure and discuss conserved and organism-specific features of Pol I.
- Florian B. Heiss
- , Julia L. Daiß
- & Christoph Engel
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Article
| Open AccessA scaffold lncRNA shapes the mitosis to meiosis switch
In fission yeast, the antagonistic RNA-binding proteins Mmi1 and Mei2 respectively promote and inhibit meiotic mRNA degradation during mitotic growth. Here the authors show that the lncRNA mamRNA scaffolds Mmi1 and Mei2 proteins to enable their mutual controls.
- Vedrana Andric
- , Alicia Nevers
- & Mathieu Rougemaille
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Article
| Open AccessNucleic acid binding by SAMHD1 contributes to the antiretroviral activity and is enhanced by the GpsN modification
SAMHD1 catalyses the dephosphorylation of deoxynucleotide triphosphates (dNTPs) and has antiretroviral activity. Here, the authors present the crystal structures of SAMHD1-oligonucleotide complexes, which reveal that the allosteric binding sites of SAMHD1 are plastic and can fit oligonucleotides in place of the two allosteric activators GTP and dNTP, and they also show that SAMHD1 recognises GpsN phosphorothioation modifications in nucleic acids, which is of interest in drug design.
- Corey H. Yu
- , Akash Bhattacharya
- & Dmitri N. Ivanov
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Article
| Open AccessCryo-EM of mammalian PA28αβ-iCP immunoproteasome reveals a distinct mechanism of proteasome activation by PA28αβ
The proteasome activator PA28αβ affects MHC class I antigen presentation by associating with immunoproteasome core particles (iCPs). Cryo-EM structures of the mammalian PA28αβ -iCP immunoproteasome and free iCP, combined with cross-linking data, reveal the complex architecture and suggest a distinct immunoproteasome activation mechanism.
- Jinhuan Chen
- , Yifan Wang
- & Yao Cong
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Article
| Open AccessMolecular mechanism for vitamin C-derived C5-glyceryl-methylcytosine DNA modification catalyzed by algal TET homologue CMD1
C5-glyceryl-methylcytosine is a DNA modification that plays a role in the regulation of green alga photosynthesis and is catalysed by CMD1, using vitamin C (VC) as a co-substrate. Here, the authors provide insights into the catalytic mechanism of CMD1 by determining the crystal structures of apo CMD1 and CMD1 bound to either VC or DNA, as well as the ternary CMD1/VC/DNA complex structure.
- Wenjing Li
- , Tianlong Zhang
- & Jianping Ding
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Article
| Open AccessStructural basis for ligand recognition of the neuropeptide Y Y2 receptor
The human neuropeptide Y receptor Y2 (Y2R) is a drug target for the treatment of obesity and anxiety. Crystal structure of Y2R bound to a selective antagonist and accompanying mutagenesis provide insights into ligand recognition and subtype specificity of NPY receptors.
- Tingting Tang
- , Christin Hartig
- & Beili Wu
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Article
| Open AccessA safety cap protects hydrogenase from oxygen attack
[FeFe]-hydrogenases catalyze the conversion of protons and electrons to molecular hydrogen, but upon exposure to oxygen, their catalytic cofactor is irreversibly inactivated. Here, the authors determine the crystal structure of hydrogenase CbA5H and identify a cysteine residue, which acts as a safety cap that shields the active site from oxygen.
- Martin Winkler
- , Jifu Duan
- & Thomas Happe
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Article
| Open AccessStructure of papain-like protease from SARS-CoV-2 and its complexes with non-covalent inhibitors
The SARS-CoV-2 papain-like protease (PLpro) is of interest as an antiviral drug target. Here, the authors synthesize and characterise naphthalene-based inhibitors for PLpro and present the crystal structures of PLpro in its apo state and with the bound inhibitors, which is of interest for further structure-based drug design efforts.
- Jerzy Osipiuk
- , Saara-Anne Azizi
- & Andrzej Joachimiak
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Article
| Open AccessBackbone-independent NMR resonance assignments of methyl probes in large proteins
Here, the authors present Methyl Assignments Using Satisfiability (MAUS), a method for the assignment of methyl groups using raw NOE data. They use eight proteins in the 10–45 kDa size range as test cases and show that MAUS yields 100% accurate assignments at high completeness levels.
- Santrupti Nerli
- , Viviane S. De Paula
- & Nikolaos G. Sgourakis
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Article
| Open AccessInfrared nanospectroscopy reveals the molecular interaction fingerprint of an aggregation inhibitor with single Aβ42 oligomers
Our understanding of the molecular mechanisms underlying pathological protein aggregation remains incomplete. Here, single molecule infrared nanospectroscopy (AFM-IR) offers insight into the structure of Aβ42 oligomeric and fibrillar species and their interaction with an aggregation inhibitor, paving the way for single molecule drug discovery studies.
- Francesco Simone Ruggeri
- , Johnny Habchi
- & Tuomas P. J. Knowles
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Article
| Open AccessDirect control of CAR T cells through small molecule-regulated antibodies
Many next-generation antibody therapeutics have enhanced potency but the risk of adverse events. Here the authors develop a conditionally activated, single-module CAR.
- Spencer Park
- , Edward Pascua
- & Javier Chaparro-Riggers
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Article
| Open AccessDutaFabs are engineered therapeutic Fab fragments that can bind two targets simultaneously
Bispecific antibodies can bind to two distinct targets though the fusion of two different Fv regions. In this study, the authors develop DutaFabs that present two separated and independent antigen binding sites within the same Fv region, giving rise to bispecific Fab fragments.
- Roland Beckmann
- , Kristian Jensen
- & Hubert Kettenberger
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Article
| Open AccessNeissLock provides an inducible protein anhydride for covalent targeting of endogenous proteins
Covalent conjugation of endogenous protein complexes offers many opportunities for fundamental and clinical research. Based on a bacterial protein domain that forms a reactive anhydride in the presence of Ca2+, the authors here develop a system that enables the covalent capture of endogenous binding partners.
- Arne H. A. Scheu
- , Sheryl Y. T. Lim
- & Mark Howarth
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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 AccessStructures of the archaerhodopsin-3 transporter reveal that disordering of internal water networks underpins receptor sensitization
Archaerhodopsin-3 (AR3) mutants are commonly used in optogenetics for neuron silencing and membrane voltage sensing. High-resolution crystal structures show that desensitization of the AR3 photoreceptor occurs when internal hydrogen-bonded water networks are modified in response to changes in chromophore isomerization.
- Juan F. Bada Juarez
- , Peter J. Judge
- & Anthony Watts
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Article
| Open AccessAPE1 distinguishes DNA substrates in exonucleolytic cleavage by induced space-filling
The exonuclease Apurinic/apyrimidinic endonuclease 1 (APE1) is important for processing matched/mismatched terminus during DNA repair. Here the authors resolve the X-ray crystallography structures of APE1 with varied dsDNAs substrates to assay the nuclease activity.
- Tung-Chang Liu
- , Chun-Ting Lin
- & Yu-Yuan Hsiao
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Article
| Open AccessCryo-EM structures of the SARS-CoV-2 endoribonuclease Nsp15 reveal insight into nuclease specificity and dynamics
Nsp15 is a uridine specific endoribonuclease present in all coronaviruses. Here, the authors determine the cryo-EM structures of SARS-CoV-2 Nsp15 in the apo and UTP-bound states, which together with biochemical experiments, mass spectrometry and molecular dynamics simulations provide insights into the catalytic mechanism of Nsp15 and its conformational dynamics.
- Monica C. Pillon
- , Meredith N. Frazier
- & Robin E. Stanley
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Article
| Open AccessHuman antibodies targeting a Mycobacterium transporter protein mediate protection against tuberculosis
Antibody responses against Mycobacteria infection have been reported, but whether and how they impact anti-bacteria immunity in the host is unclear. Here the authors characterize human anti-Mycobacteria antibodies to find them targeting a Mycobacteria transporter protein, PstS1, show distinct interaction modes in crystal structure, and mediate protection in vitro.
- Avia Watson
- , Hao Li
- & Natalia T. Freund