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| Open AccessPIE-seq: identifying RNA-binding protein targets by dual RNA-deaminase editing and sequencing
Tracking protein-RNA interaction across cell types is challenging. Here, Ruan et al develop a dual-deaminase method called PIE-Seq, where protein targets are marked by both C-to-U and A-to-I RNA base editors, and apply it to 25 human RNA-binding proteins.
- Xiangbin Ruan
- , Kaining Hu
- & Xiaochang Zhang
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Article
| Open AccessNERNST: a genetically-encoded ratiometric non-destructive sensing tool to estimate NADP(H) redox status in bacterial, plant and animal systems
NADP(H) is a crucial cofactor, acting as a reducing agent in numerous pathways in living organisms. Here the authors report a ratiometric biosensor named NERNST, which can be used to estimate the NADP(H) redox status in bacterial, plant and animal cells and organelles.
- Pamela E. Molinari
- , Adriana R. Krapp
- & Matias D. Zurbriggen
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Article
| Open AccessThe UBX domain in UBXD1 organizes ubiquitin binding at the C-terminus of the VCP/p97 AAA-ATPase
The function of VCP/p97 AAA-ATPase cofactor UBXD1 and its UBX domain has been elusive. Here the authors show that the extended UBXD1 UBX domain is located at the p97 pore exit where it binds ubiquitin, suggesting that UBXD1 receives unfolded substrates and hands them off for down-stream processing.
- Mike Blueggel
- , Alexander Kroening
- & Christine Beuck
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Article
| Open AccessDevelopment of allosteric and selective CDK2 inhibitors for contraception with negative cooperativity to cyclin binding
Despite the therapeutic interest in targeting CDK2, developing a selective CDK2 inhibitor has been challenging. Here, the authors describe a potent and selective CDK2 inhibitor that binds an allosteric pocket, preventing activating protein partners from binding and showing potential as a contraceptive.
- Erik B. Faber
- , Luxin Sun
- & Gunda I. Georg
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Article
| Open AccessCryo-EM structure of the Saccharomyces cerevisiae Rpd3L histone deacetylase complex
The Rpd3L HDAC complex is an ancient chromatin-modifying complex found in diverse eukaryotes. Here, authors describe the cryo-EM structure of the yeast complex and show that key features are preserved in the human complex.
- Avinash B. Patel
- , Jinkang Qing
- & Yuan He
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Article
| Open AccessDiscovery and biosynthesis of tricyclic copper-binding ribosomal peptides containing histidine-to-butyrine crosslinks
Cyclic peptides are important bioactive compounds and drugs, synthesised by enzymatic side-chain macrocyclization of ribosomal peptides, which rarely involves histidine residues. Here, the authors report the discovery and biosynthesis of tricyclic lanthipeptide noursin, constrained by a tri amino acid labionin crosslink and histidine-to-butyrine crosslink, which is important for copper binding of noursin.
- Yuqing Li
- , Yeying Ma
- & Huan Wang
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Article
| Open AccessDe novo cholesterol biosynthesis in bacteria
Production of highly modified sterols, such as cholesterol, is essential to eukaryotic physiology but has not been yet reported for bacteria. Here, Lee et al. show that a marine myxobacterium produces cholesterol, and provide evidence for further downstream modifications in this and other bacterial species.
- Alysha K. Lee
- , Jeremy H. Wei
- & Paula V. Welander
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Article
| Open AccessRubisco deactivation and chloroplast electron transport rates co-limit photosynthesis above optimal leaf temperature in terrestrial plants
Photosynthesis declines at mild temperatures in terrestrial plants. Here, the authors use published data to show that decline in photosynthetic CO2 assimilation rate with rising temperatures can be accounted for by Rubisco deactivation and declines in chloroplast electron transport rate.
- Andrew P. Scafaro
- , Bradley C. Posch
- & Owen K. Atkin
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Article
| Open AccessElucidating the molecular programming of a nonlinear non-ribosomal peptide synthetase responsible for fungal siderophore biosynthesis
Fungal siderophores are biosynthesised by nonribosomal peptide synthetases (NRPSs) with highly unusual domain architectures. Here, the authors characterise cryptic programming events occurring within SidC NRPS, responsible for ferricrocin biosynthesis in Aspergillus nidulans.
- Matthew Jenner
- , Yang Hai
- & Yi Tang
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Article
| Open AccessTurn air-captured CO2 with methanol into amino acid and pyruvate in an ATP/NAD(P)H-free chemoenzymatic system
The use of gaseous and air-captured CO2 for technical biosynthesis is highly desired but challenging due to high energy demands. Here, the authors present an ATP and NAD(P)H-free chemoenzymatic system for glycine, serine, and pyruvate biosynthesis by coupling methanol with gaseous and air-captured CO2.
- Jianming Liu
- , Han Zhang
- & An-Ping Zeng
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Article
| Open AccessA general model to predict small molecule substrates of enzymes based on machine and deep learning
For many enzymes, it is unknown which primary and/or secondary reactions they catalyze. Here, the authors use machine and deep learning to develop a general model for the prediction of enzyme-small molecule substrate pairs and make the resulting model available through a webserver.
- Alexander Kroll
- , Sahasra Ranjan
- & Martin J. Lercher
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Article
| Open AccessStructural insights into cardiolipin replacement by phosphatidylglycerol in a cardiolipin-lacking yeast respiratory supercomplex
Whether anionic phospholipids required for respiratory supercomplex (SC) formation is unclear. Here authors resolve SCs from a wild type and cardiolipin-deficient yeast strain at 3.2- 3.3 Å resolution to show that cardiolipin can be replaced by phosphatidylglycerol.
- Corey F. Hryc
- , Venkata K. P. S. Mallampalli
- & William Dowhan
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Article
| Open AccessStructure of a eukaryotic cholinephosphotransferase-1 reveals mechanisms of substrate recognition and catalysis
CDP-alcohol phosphatidyltransferase (CDP-AP) is a family of membrane-embedded enzymes that synthesize phospholipids. The authors report structural and functional studies of a eukaryotic CDP-AP and reveal a catalytic center and structural fold different from these of prokaryotic homologs.
- Lie Wang
- & Ming Zhou
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Article
| Open AccessOptimal enzyme utilization suggests that concentrations and thermodynamics determine binding mechanisms and enzyme saturations
One of the main challenges hampering the development of kinetic models is the lack of kinetic parameters for many enzymatic reactions. Here, the authors introduce a framework to explore the catalytically optimal operating conditions of any complex enzyme mechanism from an evolutionary perspective.
- Asli Sahin
- , Daniel R. Weilandt
- & Vassily Hatzimanikatis
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Article
| Open AccessEngineered repeat proteins as scaffolds to assemble multi-enzyme systems for efficient cell-free biosynthesis
Multi-enzymatic cascades benefit from precise nanometric organization but achieving this using available scaffolds is challenging. Here the authors present strategy for organizing multienzymatic systems using a protein scaffold based on TRAP domains, and demonstrate improved catalytic output.
- Alba Ledesma-Fernandez
- , Susana Velasco-Lozano
- & Aitziber L. Cortajarena
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Article
| Open AccessSpecific, sensitive and quantitative protein detection by in-gel fluorescence
Recombinant proteins in complex solutions are typically detected with tag-specific antibodies in Western blots. Here, the author describes an antibody-free alternative in which tagged proteins are detected directly in polyacrylamide gels via fluorophore-labelling of the tagged protein using a ligase.
- Adrian C. D. Fuchs
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Article
| Open AccessA selective and atom-economic rearrangement of uridine by cascade biocatalysis for production of pseudouridine
Pseudouridine (Ψ), the C-nucleoside isomer of uridine, and its 1-N-methyl derivative, are incorporated in mRNA vaccines and essential for their efficiency, but difficult to synthetically access. Here, the authors report on selective and atom-economic 1N-5C rearrangement of β-d-ribosyl on uracil to obtain Ψ from unprotected U in quantitative yield
- Martin Pfeiffer
- , Andrej Ribar
- & Bernd Nidetzky
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Article
| Open AccessDiscovery and structural characterization of monkeypox virus methyltransferase VP39 inhibitors reveal similarities to SARS-CoV-2 nsp14 methyltransferase
Monkeypox virus is a pathogen with pandemic potential, encoding for its own RNA capping machinery. Here, the authors present crystal structures of its 2′-O-RNA methyltransferase VP39 in complex with sub-micromolar inhibitors and reveal similarities to SARS-CoV−2 nsp14 methyltransferase.
- Jan Silhan
- , Martin Klima
- & Evzen Boura
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Article
| Open AccessIdentification of d-arabinan-degrading enzymes in mycobacteria
Bacterial cell growth and division require the coordinated action of enzymes that synthesize and degrade cell wall polymers. Here, the authors identify enzymes that cleave the D-arabinan core of arabinogalactan, an unusual component of the cell wall of Mycobacterium tuberculosis and other mycobacteria.
- Omar Al-Jourani
- , Samuel T. Benedict
- & Patrick J. Moynihan
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Article
| Open AccessStructure and mechanism of the alkane-oxidizing enzyme AlkB
Alkane monooxygenase (AlkB) plays a key role in the global carbon cycle and remediation of oil spills. Here, the authors report the cryo-EM structure of AlkB to provide insight into the catalytic mechanism and substrate selectivity.
- Xue Guo
- , Jianxiu Zhang
- & Liang Feng
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Article
| Open AccessActive and stable alcohol dehydrogenase-assembled hydrogels via synergistic bridging of triazoles and metal ions
Biocatalysis using immobilized enzymes is increasingly replacing traditional methods of manufacturing fine chemicals but heir industrial applications are limited by the low specific activity and poor stability. Here, the authors report a feasible strategy utilizing the synergistic bridging of triazoles and metal ions to induce the formation of porous enzyme-assembled hydrogels with increased activity.
- Qiang Chen
- , Ge Qu
- & Guangsheng Luo
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Article
| Open AccessExtended DNA threading through a dual-engine motor module of the activating signal co-integrator 1 complex
ASCC3 is a multi-functional helicase that contains two consecutive Ski2-like helicase units. Here, the authors show that ASCC3 can unwind DNA by threading one strand of a substrate duplex through both helicase units, supported by the TRIP4 protein.
- Junqiao Jia
- , Tarek Hilal
- & Markus C. Wahl
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Article
| Open AccessInhibitor-3 inhibits Protein Phosphatase 1 via a metal binding dynamic protein–protein interaction
Protein phosphatase 1 (PP1) is regulated by intrinsically disordered proteins like inhibitor-3, I3. The authors show that I3 does not inhibit PP1 by forming a rigid complex but instead by binding dynamically with its active site metals, showing how flexibility is used in biology.
- Gautam Srivastava
- , Meng S. Choy
- & Wolfgang Peti
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Article
| Open AccessTurning universal O into rare Bombay type blood
People with the rare Bombay-type Oh blood group can only be transfused with Oh blood. Here, the authors characterize a bacterial α−1,2-fucosidase that can convert universal O type into rare Bombay type blood.
- Itxaso Anso
- , Andreas Naegeli
- & Marcelo E. Guerin
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Article
| Open AccessMechanism of antibody-specific deglycosylation and immune evasion by Streptococcal IgG-specific endoglycosidases
Bacterial pathogens have evolved intricate mechanisms to evade the human immune system, including the production of immunomodulatory enzymes. Here, the authors establish the mechanisms of recognition and specific deglycosylation of IgG antibodies by the multi-modular enzymes EndoS and EndoS2
- Beatriz Trastoy
- , Jonathan J. Du
- & Marcelo E. Guerin
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Article
| Open AccessUnnatural activities and mechanistic insights of cytochrome P450 PikC gained from site-specific mutagenesis by non-canonical amino acids
The 20 proteinogenic amino acids cannot always satisfy the requirements of protein engineering. Here, the authors practice semi-rational non-canonical amino acid mutagenesis for a cytochrome P450 enzyme, leading to unnatural activities and significant mechanistic insights.
- Yunjun Pan
- , Guobang Li
- & Shengying Li
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Article
| Open AccessNeutron-encoded diubiquitins to profile linkage selectivity of deubiquitinating enzymes
Most insights into deubiquitinase (DUB) substrate specificity originate from studies with isolated di-ubiquitins (diUb), but in cells diUbs with different linkage types coexist. Here, the authors develop a mass spectrometric DUB activity assay that can probe all diUbs simultaneously under substrate competition conditions.
- Bianca D. M. van Tol
- , Bjorn R. van Doodewaerd
- & Paul P. Geurink
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Article
| Open AccessComplete bio-degradation of poly(butylene adipate-co-terephthalate) via engineered cutinases
Bio-degradation of poly(butylene adipate-co-terephthalate) is an attractive tactic but requires an effective hydrolytic enzyme. Here, the authors demonstrate that cutinases are highly potent PBAT-decomposing enzymes and their mechanism is proposed based on substrate-binding mode.
- Yu Yang
- , Jian Min
- & Rey-Ting Guo
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Article
| Open AccessGenome mining unveils a class of ribosomal peptides with two amino termini
RiPP discovery has expanded the scope of post-translational modification chemistry, but genome mining of RiPP classes remains an unsolved challenge. Here, the authors employed bioinformatics and synthetic biology approaches to discover and characterize an unknown class of RiPPs, defined by an unusual amino-modified C-terminus.
- Hengqian Ren
- , Shravan R. Dommaraju
- & Huimin Zhao
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Article
| Open AccessRetinol dehydrogenase 10 reduction mediated retinol metabolism disorder promotes diabetic cardiomyopathy in male mice
The current challenges for diabetic cardiomyopathy (DCM) are unclear mechanisms and no effective therapy in clinics. Here, the authors found that the decrease of cardiac retinol dehydrogenase 10 in type 2 diabetes leads to retinol metabolism disorder, cardiac lipid toxicity and cardiomyopathy development, suggesting that correcting the imbalance of cardiac retinol metabolism may be an effective strategy for the treatment of DCM.
- Yandi Wu
- , Tongsheng Huang
- & Weibin Cai
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Article
| Open AccessEvolution of enzyme functionality in the flavin-containing monooxygenases
Detoxification enzymes are crucial for the survival of animals in new environments. Here, the authors study the molecular mechanism behind the catalytic diversification of a major family of tetrapod detoxification enzymes—the FMOs—during evolution.
- Gautier Bailleul
- , Guang Yang
- & Maria Laura Mascotti
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Article
| Open AccessVisible light-exposed lignin facilitates cellulose solubilization by lytic polysaccharide monooxygenases
Degradation of plant biomass, comprised of cellulose and polyaromatic lignin, is promoted by light. Here, the authors show that light promotes lignin-catalyzed generation of hydrogen peroxide, which is used by redox enzymes to degrade cellulose.
- Eirik G. Kommedal
- , Camilla F. Angeltveit
- & Vincent G. H. Eijsink
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Article
| Open AccessStructural remodelling of the carbon–phosphorus lyase machinery by a dual ABC ATPase
Here, authors analyse the structural organisation of the large carbon-phosphorus lyase enzyme from bacteria using electron microscopy and discover that it contains two ATP-binding cassette dimers of PhnK and PhnL and opens upon ATP hydrolysis.
- Søren K. Amstrup
- , Sui Ching Ong
- & Ditlev E. Brodersen
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Article
| Open AccessGenomics and biochemical analyses reveal a metabolon key to β-L-ODAP biosynthesis in Lathyrus sativus
Grass pea is a multi-stress tolerant orphan crop and developing cultivars with decreased accumulation of the neurotoxin β-L-oxalyl-2,3-diaminopropionic acid (β-L-ODAP) is one of its breeding objectives. Here, the authors assemble its genome and reveal genes involved in the biosynthesis of β-L-ODAP.
- Anne Edwards
- , Isaac Njaci
- & Peter M. F. Emmrich
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Article
| Open AccessBacterial origins of thymidylate metabolism in Asgard archaea and Eukarya
Asgard archaea include the closest known archaeal relatives of eukaryotes. Here, the authors provide evidence that eukaryotic and Asgard thymidylate synthases (required for DNA synthesis) may have a bacterial origin, and additional lateral transfer of bacterial genes may have shaped the metabolism of Asgard archaea.
- Jonathan Filée
- , Hubert F. Becker
- & Hannu Myllykallio
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Article
| Open AccessA genome-wide CRISPR-Cas9 knockout screen identifies FSP1 as the warfarin-resistant vitamin K reductase
The authors develop a Vitamin K-dependent apoptotic reporter cell line for large-scale screening of enzymes associated with Vitamin K-dependent carboxylation and identify ferroptosis suppressor protein 1 (FSP1) as responsible for warfarin-resistant vitamin K reduction.
- Da-Yun Jin
- , Xuejie Chen
- & Jian-Ke Tie
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Article
| Open AccessEmergence of a proton exchange-based isomerization and lactonization mechanism in the plant coumarin synthase COSY
Plant biosynthetic enzymes rapidly evolve to catalyze specialized reactions. Here, the authors present the crystal structure and mechanism of COSY, the enzyme involved in coumarin biosynthesis of the BAHD-acyltransferase family that catalyzes an intramolecular acyl transfer reaction through a proton exchange mechanism.
- Colin Y. Kim
- , Andrew J. Mitchell
- & Jing-Ke Weng
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Article
| Open AccessA paralog of Pcc1 is the fifth core subunit of the KEOPS tRNA-modifying complex in Archaea
Many eukaryotic and archaeal tRNAs carry a modified adenosine (t6A) that is synthesized by the KEOPS complex, which is composed of four subunits. A fifth subunit (Gon7) is found only in fungi and metazoa. Here the authors show that archaea also possess a fifth subunit, which is structurally and functionally similar to eukaryotic Gon7.
- Marie-Claire Daugeron
- , Sophia Missoury
- & Tamara Basta
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Article
| Open AccessIron-sulfur clusters are involved in post-translational arginylation
The enzyme ATE1 catalyzes eukaryotic post-translation arginylation, a key protein modification necessary for cellular homeostasis. Here, the authors show that ATE1s are previously unrealized iron-sulfur proteins that use this oxygen-sensitive inorganic cofactor to control cellular arginylation
- Verna Van
- , Janae B. Brown
- & Aaron T. Smith
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Article
| Open AccessThe unifying catalytic mechanism of the RING-between-RING E3 ubiquitin ligase family
RBR E3 ubiquitin ligases utilise a 2-step catalytic mechanism previously defined for only few of the RBR family members. Here, the authors examine the poorly studied RBRs HOIL-1 and RNF216 to define general principles of RBR catalysis and regulation and identify specific functional differences.
- Xiangyi S. Wang
- , Thomas R. Cotton
- & Bernhard C. Lechtenberg
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Article
| Open AccessSourcing thermotolerant poly(ethylene terephthalate) hydrolase scaffolds from natural diversity
Enzymes have potential for recycling plastics such as PET, a polyester used in textiles and single-use packaging. Here, the authors identify and characterize additional PET-active biocatalysts and expand the number and diversity of thermotolerant scaffolds for enzymatic PET deconstruction.
- Erika Erickson
- , Japheth E. Gado
- & John E. McGeehan
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Article
| Open AccessRational enzyme design for enabling biocatalytic Baldwin cyclization and asymmetric synthesis of chiral heterocycles
Chiral heterocyclic compounds are privileged structures in medicinal chemistry. Here, the authors report an in silico strategy for the enzymatic synthesis of pharmaceutically significant chiral N- and O-heterocycles via Baldwin cyclization of hydroxy- and amino-substituted epoxides and oxetanes using epoxide hydrolase mutants.
- Jun-Kuan Li
- , Ge Qu
- & Zhoutong Sun
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Article
| Open AccessCompact Cas9d and HEARO enzymes for genome editing discovered from uncultivated microbes
Programmable, RNA-guided nucleases are diverse enzymes that have been repurposed for biotechnological applications. Here, the authors mine an extensive genome-resolved metagenomics database and identified uncharacterized families of RNA-guided, compact nucleases.
- Daniela S. Aliaga Goltsman
- , Lisa M. Alexander
- & Christopher T. Brown
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Article
| Open AccessStructure of the AlgKX modification and secretion complex required for alginate production and biofilm attachment in Pseudomonas aeruginosa
The pathogen Pseudomonas aeruginosa uses a synthase-dependent secretion system for production of the exopolysaccharide alginate, which is associated with lung infection severity. Here, Gheorghita et al. determine the crystal structure of one of the secretion system components (the AlgKX complex) and show that it binds alginate oligosaccharides and is required for polymer production and biofilm attachment.
- Andreea A. Gheorghita
- , Yancheng E. Li
- & P. Lynne Howell
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Article
| Open AccessStructural basis for the non-self RNA-activated protease activity of the type III-E CRISPR nuclease-protease Craspase
The authors report several high-resolution functional snapshots of type III-E nuclease-protease Craspase complexes, revealing the mechanisms underlying target RNA cleavage and non-self RNA activated protease activities; and highlighting the potentials for the development of RNA-guided nuclease-protease Craspase-based tools for biotechnological applications.
- Ning Cui
- , Jun-Tao Zhang
- & Ning Jia
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Article
| Open AccessSESAME-catalyzed H3T11 phosphorylation inhibits Dot1-catalyzed H3K79me3 to regulate autophagy and telomere silencing
How H3T11 phosphorylation exerts biological functions remains poorly understood. Here, authors show that H3pT11 directly inhibits Dot1-catalyzed H3K79 tri-methylation (H3K79me3) and uncover how this histone crosstalk regulates autophagy and telomere silencing.
- Fei He
- , Qi Yu
- & Shanshan Li
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Article
| Open AccessA growth selection system for the directed evolution of amine-forming or converting enzymes
Fast screening of enzymes is key for directed evolution of industrial biocatalysts. Here, the authors report a simple, high-throughput, and low-equipment-dependent growth selection system for engineering three enzymes for synthesis of chiral amines.
- Shuke Wu
- , Chao Xiang
- & Uwe T. Bornscheuer
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Article
| Open AccessStructural insights into mechanism and specificity of the plant protein O-fucosyltransferase SPINDLY
Arabidopsis SPINDLY (SPY) is a nucleocytoplasmic protein O-fucosyltransferase. Here, the authors present a crystal structure of Arabidopsis SPY/GDP complex, reveal SPY’s substrate recognition and enzyme mechanism, and provide insights into the glycan donor substrate selection in GT41 proteins.
- Li Zhu
- , Xiting Wei
- & Shutong Xu
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Article
| Open AccessTotal enzymatic synthesis of cis-α-irone from a simple carbon source
Retrosynthetic pathway design using promiscuous enzymes can provide a solution to the biosynthetic production of natural products. Here, the authors design a pathway for the production of cis-α-irone with a promiscuous methyltransferase using structure-guided enzyme engineering strategies.
- Xixian Chen
- , Rehka T
- & Isabelle André