Biochemistry

  • Article
    | Open Access

    The polycomb repressive complex 2 (PRC2) is a histone methyltransferase regulating cell differentiation and identity. Here, the authors show that the vertebrate-specific PRC2 accessory subunit PALI1 facilitates substrate binding by the complex and elucidate the allosteric mechanism of PALI1- mediated PRC2 activation.

    • Qi Zhang
    • , Samuel C. Agius
    •  & Chen Davidovich
  • Article
    | Open Access

    Mitochondrial transport toward both the plus- and minus-ends of microtubules is mediated by motor proteins linked to mitochondria by TRAK adaptor proteins. Here the authors investigate the role of TRAK2 as a bidirectional motor adaptor, and propose a model where TRAK2 coordinates the activities of opposing kinesin-1 and cytoplasmic dynein motors as a single interdependent motor complex.

    • Adam R. Fenton
    • , Thomas A. Jongens
    •  & Erika L. F. Holzbaur
  • Article
    | Open Access

    PARG and ARH3 are the main hydrolases to reverse serine poly(ADP-ribosylation) yet their activities in the process differ. Here, the authors synthesise linear and branched poly(ADP-ribose) molecules, perform structure-function analysis and elucidate the mechanistic differences between PARG and ARH3.

    • Johannes Gregor Matthias Rack
    • , Qiang Liu
    •  & Ivan Ahel
  • Article
    | Open Access

    The bacterial thymidylate synthase ThyX catalyzes the reductive methylation of deoxyuridylate (dUMP) into deoxythymidylate (dTMP) and requires both folate and flavin for activity. Here, the authors combine biochemical experiments, spectroscopic measurements and flavin synthesis chemistry to show that formaldehyde (CH2O) can replace the natural methylene donor of ThyX in a CH2O-shunt reaction, yielding a carbinolamine intermediate with the reduced flavin coenzyme, and they present the crystal structure of this intermediate.

    • Charles Bou-Nader
    • , Frederick W. Stull
    •  & Djemel Hamdane
  • Article
    | Open Access

    Assembly of the mitoribosome requires assistance from numerous specialized factors. Here, structures of the human 39S late assembly intermediates identify several assembly factors which keep the 16S rRNA in immature conformations, and reveal deacylated tRNA in the ribosomal E-site, suggesting a role in 39S assembly.

    • Jingdong Cheng
    • , Otto Berninghausen
    •  & Roland Beckmann
  • Article
    | Open Access

    Human cytomegalovirus (HCMV) is the prototypical member of the β-herpesvirinae subfamily and the leading viral cause of congenital infections that can lead to birth defects and it can also cause life-threatening disease in immunocompromised individuals. Here, the authors present the in-situ cryo-EM structures of the symmetry-mismatched portal and the capsid vertex-specific components (CVSCs) of HCMV and discuss the mechanistic implications for genome package, retention and ejection.

    • Zhihai Li
    • , Jingjing Pang
    •  & Xuekui Yu
  • Article
    | Open Access

    Nutrient status in the cell regulates autophagy via mTORC1 activity. Here, the authors show that the ubiquitous G protein subunit Gαq contributes to nutrient sensing by promoting formation of an mTOR-p62-Raptor complex in replete conditions, modulating autophagy.

    • Sofía Cabezudo
    • , Maria Sanz-Flores
    •  & Catalina Ribas
  • Article
    | Open Access

    The transmembrane protein Wntless (WLS) is essential for the secretion of Wnt proteins, which are signaling molecules of the Wnt signaling pathways. Here, the authors present the 2.2 Å cryo-EM structure of human WLS in complex with Wnt3a and discuss mechanistic implications for the palmitoleoylation of Wnt3a that is required for WLS mediated Wnt secretion.

    • Qing Zhong
    • , Yanyu Zhao
    •  & Dan Ma
  • Article
    | Open Access

    Pannexin 1 (PANX1) is a membrane channel mediating release of signaling molecules to the extracellular space. PANX1 can be activated by GPCRs. Here, the authors elucidate a non-canonical channel activation pathway by α1-adrenergic receptor that involves HDAC6- mediated lysine deacetylation of PANX1.

    • Yu-Hsin Chiu
    • , Christopher B. Medina
    •  & Douglas A. Bayliss
  • Article
    | Open Access

    The nucleation mechanisms of biological protein phase separation are poorly understood. Here, the authors perform time-resolved SAXS experiments with the low-complexity domain (LCD) of hnRNPA1 and uncover multiple kinetic regimes on the micro- to millisecond timescale. Initially, individual proteins collapse. Nucleation then occurs via two steps distinguished by their protein cluster size distributions.

    • Erik W. Martin
    • , Tyler S. Harmon
    •  & Tanja Mittag
  • Article
    | Open Access

    Calcium signals initiated by IP3 receptors in ER membranes regulate most cellular activities. Here, the authors show that KRas-induced actininteracting protein (KRAP) tethers a small subset of IP3 receptors to actin and licenses them to evoke cytosolic calcium signals.

    • Nagendra Babu Thillaiappan
    • , Holly A. Smith
    •  & Colin W. Taylor
  • Article
    | Open Access

    Self-assembling peptides have a range of potential applications but developing self-assembling sequences can be challenging. Here, the authors report on a one-bead one-compound combinatorial library where fluorescence is used to detect the potential for self-assembly and identified candidates are evaluated.

    • Pei-Pei Yang
    • , Yi-Jing Li
    •  & Kit S. Lam
  • Article
    | Open Access

    The Class 2 family of CRISPR nucleases named Cas12j, which shares only low sequence identity with other CRISPR nucleases was recently identified in the biggiephage clade of phages. Here, the authors present the cryo-EM structure of a functional Cas12j3−crRNA complex in the post-catalytic state and discuss Cas12j3 PAM recognition, hybrid stabilisation and the activation mechanism.

    • Arturo Carabias
    • , Anders Fuglsang
    •  & Guillermo Montoya
  • Article
    | Open Access

    Plant sulfate transporters mediate absorption and distribution of sulfate. Here the authors present functional assays and a cryo-EM structure of the Arabidopsis SULTR4;1 transporter identifying key domains for dimerization, substrate binding and coupling of transport activity to a proton gradient.

    • Lie Wang
    • , Kehan Chen
    •  & Ming Zhou
  • Article
    | Open Access

    Currently many of the time resolved serial femtosecond (SFX) crystallography experiments are done with light driven protein systems, whereas the reaction initiation for non-light triggered enzymes remains a major bottle neck. Here, the authors present an expanded Drop-on-Tape system, where picoliter-sized droplets of a substrate or inhibitor are turbulently mixed with nanoliter sized droplets of microcrystal slurries, and they use it for time-resolved SFX measurements of inhibitor binding to lysozyme and secondly, binding of a β-lactam antibiotic to a bacterial serine β-lactamase.

    • Agata Butryn
    • , Philipp S. Simon
    •  & Allen M. Orville
  • Article
    | Open Access

    Many endoperoxide-containing natural products have been isolated, but the biosynthesis of the endoperoxides remains unclear. Here, the authors report the structural and functional analysis of the NvfI endoperoxidase that catalyzes the formation of fumigatonoid A in the biosynthesis of novofumigatonin, and show that it does not employ tyrosyl radical in the reaction.

    • Takahiro Mori
    • , Rui Zhai
    •  & Ikuro Abe
  • Article
    | Open Access

    The bacteriophytochrome DrBphP from Deinococcus radiodurans shows high sequence homology to the histidine kinase Agp1 from Agrobacterium fabrum but lacks kinase activity. Here, the authors structurally and biochemically analyse DrBphP and Agp1, showing that DrBphP is a light-activatable phosphatase.

    • Elina Multamäki
    • , Rahul Nanekar
    •  & Heikki Takala
  • Article
    | Open Access

    DNA probes used in next generation sequencing (NGS) have variable hybridisation kinetics, resulting in non-uniform coverage. Here, the authors develop a deep learning model to predict NGS depth using DNA probe sequences and apply to human and non-human sequencing panels.

    • Jinny X. Zhang
    • , Boyan Yordanov
    •  & David Yu Zhang
  • Article
    | Open Access

    HAP2 is essential for gamete fusion during fertilization and is conserved among eukaryotes. Here the authors show that species-specific adhesion between Chlamydomonas plus and minus gametes initiates HAP2 to undergo a fusogenic conformational change into homotrimers via a molecular mechanism akin to that of enveloped viruses.

    • Jun Zhang
    • , Jennifer F. Pinello
    •  & William J. Snell
  • Article
    | Open Access

    Histone H3K9 methylation (H3K9me) states define repressed chromatin in eukaryotic cells. Here the authors reveal complete loss of all H3K9me in mammalian cells through successive deletion of H3K9 methyltransferase genes that results in the dissolution of heterochromatin and the derepression of nearly all repeat families.

    • Thomas Montavon
    • , Nicholas Shukeir
    •  & Thomas Jenuwein
  • Article
    | Open Access

    The small molecule DNMDP acts as a velcrin by inducing complex formation between phosphodiesterase PDE3A and SLFN12, which kills cancer cells that express sufficient levels of both proteins. Here, the authors present the cryo-EM structure of the DNMDP-stabilized PDE3A-SLFN12 complex and show that SLFN12 is an RNase. PDE3A binding increases SLFN12 RNase activity, and SLFN12 RNase activity is required for DNMDP-mediated cancer cell killing.

    • Colin W. Garvie
    • , Xiaoyun Wu
    •  & Heidi Greulich
  • Article
    | Open Access

    Pleckstrin homology (PH) domains are presumed to bind phosphoinositides (PIPs) but only few specific interactions are known. Using TIRF microscopy based assays, functional studies and an experimentally validated prediction algorithm, the authors show that specific PIP binding is widespread among human PH domains.

    • Nilmani Singh
    • , Adriana Reyes-Ordoñez
    •  & Jie Chen
  • Article
    | Open Access

    There is a need to optimise cryo-EM data acquisition approaches to improve the resolution of GPCR cryo-EM structures to better than 2.5 Å, in order to use them for structure-based drug design purposes. Here, the authors present a systematic analysis of the main cryo-EM experimental parameters using three GPCRs as test cases, which is also of interest for the cryo-EM structure determination of other small membrane proteins.

    • Radostin Danev
    • , Matthew Belousoff
    •  & Patrick M. Sexton
  • Article
    | Open Access

    Reactive oxygen species are required for the long lifespan, and glutathione is an antioxidant. Here the authors show that limiting the consumption of dietary thiols, including those naturally derived from the microbiota, increases proteotoxic stress resistance in worms and human cells, and extends C. elegans lifespan.

    • Ivan Gusarov
    • , Ilya Shamovsky
    •  & Evgeny Nudler
  • Article
    | Open Access

    Noroviruses can cause gastroenteritis and there is currently no licensed vaccine or specific treatment available. Here, the authors isolate human monoclonal antibodies and characterize one antibody (NORO-320) with broad reactivity to genogroup I and II noroviruses.

    • Gabriela Alvarado
    • , Wilhelm Salmen
    •  & James E. Crowe Jr.
  • Article
    | Open Access

    Fly Dicer-2 is thought to use two distinct – processive or distributive – modes of cleavage by distinguishing the terminal structures of double-stranded RNA (dsRNA) substrates with the help of its cofactor LoquaciousPD (Loqs-PD). Here the authors show by single-molecule imaging that dsRNA terminal structures and Loqs-PD change the probability for Dicer to initiate processive cleavage but not the mode of cleavage action per se.

    • Masahiro Naganuma
    • , Hisashi Tadakuma
    •  & Yukihide Tomari
  • Article
    | Open Access

    Huntingtin exon-1 (HTTex1) consists of a N-terminal N17 domain, the disease causing polyQ domain and a C-terminal proline-rich domain (PRD). Here, the authors combine electron paramagnetic resonance (EPR), solid-state NMR with other biophysical method to characterise the structural differences of various HTTex1 fibril types with different toxicity and find that the dynamics and entanglement of the PRD domain differs among them and that the HTTex1 fibrils can be interconverted.

    • J. Mario Isas
    • , Nitin K. Pandey
    •  & Ansgar B. Siemer
  • Article
    | Open Access

    Finding durable, high-density media for data storage is necessary to support the ever-expanding generation of digital data. Here, the authors use peptide sequences to store digital data and retrieve them using tandem mass spectrometry, proving that peptides can be used as a storage medium.

    • Cheuk Chi A. Ng
    • , Wai Man Tam
    •  & Zhong-Ping Yao
  • Article
    | Open Access

    In Gram-positive bacteria, lipoprotein intramolecular transacylase Lit produces a lipoprotein variant with less immunogenicity. As such, Lit can be viewed as a virulence factor. Here, structural and functional characterization of the enzyme provides insight into its catalytic mechanism, setting the stage for future studies of Lit as a target for new antibiotics.

    • Samir Olatunji
    • , Katherine Bowen
    •  & Martin Caffrey
  • Article
    | Open Access

    Charting the landscape of 5hmC in human tissues is fundamental to understanding its regulatory functions. Here, we systematically profiled the whole-genome 5hmC landscape at single-base resolution for 19 types of human tissues and found 5hmC shows tissue-specific patterns.

    • Bo He
    • , Chao Zhang
    •  & Chengqi Yi
  • Article
    | Open Access

    Tapasin is part of the peptide loading complex necessary for presenting antigenic peptides on MHC-I for the induction of adaptive immunity. Here the authors show that tapasin interacts with MHC-I in both conserved and allele-specific regions to promote antigen presentation, with tapasin L18 and K16 residues both implicated in this molecular interaction.

    • Huan Lan
    • , Esam T. Abualrous
    •  & Christian Freund
  • Article
    | Open Access

    The authors present a method for the conversion of full-length tau protein into seeding-competent amyloid fibrils without heparin or other negatively charged co-factors, which could be useful for studying the effects of post-translational modifications on Tau aggregation as well as to identify potential inhibitors of tau aggregation. Biochemical experiments and solid-state NMR spectroscopy measurements show that these co-factor-free tau fibrils have similar properties as amyloid fibrils isolated from brain tissue but differ from those of commonly used heparin-induced tau fibrils.

    • Pijush Chakraborty
    • , Gwladys Rivière
    •  & Markus Zweckstetter
  • Article
    | Open Access

    An increase in glycine decarboxylase (GLDC) activity, a key enzyme for glycine catabolism, has been associated to tumourigenesis. Here, the authors show that mTORC1 activation induces GLDC deacetylation which impairs its ubiquitin-associated degradation leading to increased GLDC activity and tumourigenesis.

    • Rui Liu
    • , Lin-Wen Zeng
    •  & Shu Li
  • Article
    | Open Access

    The folding of outer membrane proteins (OMPs) is catalyzed by the βbarrel assembly machinery (BAM). Here, structural and functional analyses of BAM stabilized in distinct conformations elucidate the roles of lateral gate opening and interactions of BAM with the lipid bilayer in OMP assembly.

    • Paul White
    • , Samuel F. Haysom
    •  & Sheena E. Radford
  • Article
    | Open Access

    Mammalian reovirus (MRV) is a double-stranded RNA (dsRNA) virus that affects the gastrointestinal and respiratory tracts. Here, the authors present the 3.3 Å cryo-EM asymmetric reconstruction of transcribing MRV that reveals the organization of the dsRNA genome, RNA interaction with the polymerase complex, and how the polymerase interacts extensively with its co-factor, µ2, to form a transcription enzyme complex, which engages and regulates RNA transcription.

    • Muchen Pan
    • , Ana L. Alvarez-Cabrera
    •  & Z. Hong Zhou
  • Article
    | Open Access

    Prenylated indole alkaloids contain spirooxindole rings with a 3R or 3S carbon stereocenter, which determines their bioactivities, but the biocatalytic mechanism controlling the 3R- or 3S-spirooxindole formation was unclear. Here, the authors report the biochemical and structural characterization of the oxygenase/semipinacolase CtdE that catalyses the 3S-spirooxindole construction in the biosynthesis of 21R-citrinadin A.

    • Zhiwen Liu
    • , Fanglong Zhao
    •  & Xue Gao
  • Article
    | Open Access

    The fate of soil carbon is controlled by plant inputs, microbial activity, and the soil matrix. Here the authors extend the notion of plant-derived particulate organic matter, from an easily available and labile carbon substrate, to a functional component at which persistence of soil carbon is determined.

    • Kristina Witzgall
    • , Alix Vidal
    •  & Carsten W. Mueller
  • Article
    | Open Access

    The cryo-EM structure of pentameric green-light absorbing proteorhodopsin together with molecular dynamics simulations and functional studies provides insights into the proton translocation pathway and oligomerization, and a protonation-dependent mechanism for intracellular half channel hydration.

    • Stephan Hirschi
    • , David Kalbermatter
    •  & Dimitrios Fotiadis
  • Article
    | Open Access

    Chromosomal translocations involving the AF10 gene, especially with CALM, are associated with aggressive leukemias. Here the authors show that the PZP domain of AF10, a histone reader, is always excluded/impaired in AF10 fusions, whereas incorporation of this domain downregulates Hoxa genes and blocks leukemogenesis.

    • Brianna J. Klein
    • , Anagha Deshpande
    •  & Tatiana G. Kutateladze
  • Article
    | Open Access

    hNEIL1 (human endonuclease VIII-like 1) is a broadly specific DNA glycosylase for base excision repair. Here, the authors show that hNEIL1 can assume activated or triage conformations: the structural basis for the mechanism that enables broad specificity and reduces futile repair of normal bases.

    • Menghao Liu
    • , Jun Zhang
    •  & Chengqi Yi
  • Article
    | Open Access

    Junin virus (JUNV) causes Argentine hemorrhagic fever and encodes the large protein (L) of the RNA dependent RNA polymerase (RdRp) and its regulator, the matrix zinc-binding protein (Z). Here, the authors present the 3.54 Å cryo-EM structure of the complex of JUNV L with Z, and they propose a model of how JUNV L is regulated by Z during the viral life cycle and RNA synthesis.

    • Huiling Kang
    • , Jingyuan Cong
    •  & Zihe Rao
  • Article
    | Open Access

    α-ketoglutarate (αKG) is an intermediate in the tricarboxylic acid cycle that is required in the nucleus for genomic DNA demethylation by Tet3. Here, the authors show that the enzyme glutamate dehydrogenase, which converts glutamate to αKG, is redirected from the mitochondria to the nucleus.

    • Franziska R. Traube
    • , Dilara Özdemir
    •  & Thomas Carell
  • Article
    | Open Access

    The intracellular compartment is a crowded environment. Here, the authors use molecular dynamics (MD) simulations to assess inhibitor binding to c-Src kinase and show how ligand binding pathways differ in crowded and dilute protein solutions, highlighting the role of c-Src Tyr82 sidechain.

    • Kento Kasahara
    • , Suyong Re
    •  & Yuji Sugita
  • Article
    | Open Access

    Mucins play critical roles in maintaining the human microbiome, with their O-glycosylated tandem repeats (TRs) providing important cues for microbiota. Here, the authors develop a cellular platform for producing TRs with defined O-glycan structures to dissect the functions of TR O-glycosylation.

    • Rebecca Nason
    • , Christian Büll
    •  & Yoshiki Narimatsu