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| 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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| 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
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Article
| Open AccessStructure- and computational-aided engineering of an oxidase to produce isoeugenol from a lignin-derived compound
Lignin can be depolymerized into 4-alkylphenols by chemical means. Here the authors show a three-step computational-assisted enzyme engineering process to generate a biocatalyst for the conversion of lignin-derived 4-n-propylguaiacol into isoeugenol, a valuable compound.
- Yiming Guo
- , Laura Alvigini
- & Marco W. Fraaije
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Article
| Open AccessBiodegradation of highly crystallized poly(ethylene terephthalate) through cell surface codisplay of bacterial PETase and hydrophobin
High-crystallinity poly(ethylene terephthalate) is a major recycling challenge. Here, the authors show an engineered whole-cell biocatalyst showing adhesive hydrophobin and PETase on the surface of cells, for biodegradation of PET.
- Zhuozhi Chen
- , Rongdi Duan
- & Zefang Wang
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Article
| Open AccessUnscheduled DNA replication in G1 causes genome instability and damage signatures indicative of replication collisions
Reusswig et al. use engineered systems to force DNA replication in the G1 phase of the cell cycle. This unscheduled G1 replication shows hallmarks of S phase replication, but leads to over-replication and DNA breaks from replication collisions.
- Karl-Uwe Reusswig
- , Julia Bittmann
- & Boris Pfander
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Article
| Open AccessH2A.Z deposition by SWR1C involves multiple ATP-dependent steps
The multi-subunit SWR1C remodeler deposits histone variant H2A.Z at nucleosomes flanking protein-coding genes. Here the authors use single-molecule and ensemble methodologies to identify three ATP-dependent phases in the H2A.Z deposition reaction.
- Jiayi Fan
- , Andrew T. Moreno
- & Craig L. Peterson
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Article
| Open AccessXNAzymes targeting the SARS-CoV-2 genome inhibit viral infection
RNA viruses have been responsible for large-scale epidemics and pandemics throughout the last few centuries. Here, the authors show the design, synthesis and screening of artificial RNA endonuclease XNAzymes capable of cleaving genomic SARS-CoV-2 RNA and self-assembling into enzymatic nanostructures inhibiting cellular viral replication.
- Pehuén Pereyra Gerber
- , Maria J. Donde
- & Alexander I. Taylor
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Article
| Open AccessAnti-infective therapy using species-specific activators of Staphylococcus aureus ClpP
The development of selective ClpP activators targeting only the MRSA isolates without interfering with the human variant is currently challenging. Here, the authors report on the structure-based design of enantiomers of ZG197 and identify the discriminator factor between the proteins.
- Bingyan Wei
- , Tao Zhang
- & Cai-Guang Yang
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Article
| Open AccessCryo-EM structures reveal the activation and substrate recognition mechanism of human enteropeptidase
Dysfunction of enteropeptidase (EP) can result in pancreatitis, a life-threatening disease. Yang et al. report a series of EP structures representing the EP catalytic cycle and propose a mechanistic model of enzyme function.
- Xiaoli Yang
- , Zhanyu Ding
- & Haojie Huang
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Article
| Open AccessLipid-mediated activation of plasma membrane-localized deubiquitylating enzymes modulate endosomal trafficking
The endocytic degradation of plasma membrane proteins can be modulated by deubiquitylating enzymes (DUBs). Here, the authors describe two plasma membrane localized Arabidopsis DUBs that can be activated by binding to anionic lipids and influence the endocytic transport of plasma membrane proteins.
- Karin Vogel
- , Tobias Bläske
- & Erika Isono
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Article
| Open AccessDesign and directed evolution of noncanonical β-stereoselective metalloglycosidases
Metallohydrolases are ubiquitous enzymes. Here the authors show the design and biochemical characterisation of metalloglycosidase by constructing a hydrolytically active Zn-binding site within a barrel-shaped outer membrane protein OmpF.
- Woo Jae Jeong
- & Woon Ju Song
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Article
| Open AccessNicking mechanism underlying the DNA phosphorothioate-sensing antiphage defense by SspE
SspABCD–SspE is a phosphorothioation-sensing bacterial defence system. Here, authors find that SspE exploits DNA phosphorothioation-binding preference to modulate its biochemical activities for the self/nonself discrimination and phage resistance.
- Haiyan Gao
- , Xinqi Gong
- & Lianrong Wang
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Article
| Open AccessAn active site loop toggles between conformations to control antibiotic hydrolysis and inhibition potency for CTX-M β-lactamase drug-resistance enzymes
β-lactamases are a source of antibiotic resistance. Here, the authors show that an active site loop in β-lactamase toggles between conformations that control antibiotic hydrolysis and inhibitor susceptibility, highlighting the role of active site conformations in enzyme activity and inhibition.
- Shuo Lu
- , Liya Hu
- & Timothy Palzkill
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Article
| Open AccessEndowing homodimeric carbamoyltransferase GdmN with iterative functions through structural characterization and mechanistic studies
Carbamoyltransferases are a class of enzymes catalyzing carbamoylation of primary and secondary metabolites. Here, the authors show the molecular structure of the ansamycin-modifying carbamoyltransferase GdmN and repurpose the enzyme for the iterative carbamoylation of ansamitocins.
- Jianhua Wei
- , Xuan Zhang
- & Linquan Bai
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Article
| Open AccessCooperative assembly of p97 complexes involved in replication termination
This study describes how p97Ufd1-Npl4 and the UBA-UBX protein Ubxn7 disassemble vertebrate replisomes during replication termination, and it provides novel insights into how p97 complexes assemble with UBA-UBX proteins on ubiquitylated substrates
- Olga V. Kochenova
- , Sirisha Mukkavalli
- & Johannes C. Walter
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Article
| Open AccessUnexpected assembly machinery for 4(3H)-quinazolinone scaffold synthesis
4(3H)-quinazolinone is the core scaffold in more than 200 natural alkaloids and numerous drugs. Here, the authors show an alternative assembly machinery for 4(3H)-quinazolinone mainly includes a two-module NRPS catalysing tripeptide formation, unusual N-3 original sources and an α-KGD catalysing the C-N bond oxidative cleavage of a tripeptide to form a dipeptide.
- Xi-Wei Chen
- , Li Rao
- & Yi Zou
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Article
| Open AccessEndothelial alpha globin is a nitrite reductase
In mammals, hypoxia causes dilation of small arteries for increased metabolic demand. Keller et al used novel transgenic mice to show alpha hemoglobin in endothelium, once thought only in red blood cells, can regulate hypoxic-mediated dilation.
- T. C. Stevenson Keller IV
- , Christophe Lechauve
- & Brant E. Isakson
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Article
| Open AccessA universal glycoenzyme biosynthesis pipeline that enables efficient cell-free remodeling of glycans
Access to glycoenzymes for basic and applied research is limited by difficulties with their recombinant expression. Here, the authors describe a universal strategy for converting membrane-bound glycosyltransferases into water-soluble biocatalysts, which are expressed at high levels with retention of activity.
- Thapakorn Jaroentomeechai
- , Yong Hyun Kwon
- & Matthew P. DeLisa
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Article
| Open AccessMolecular basis for isoform-selective inhibition of presenilin-1 by MRK-560
Presenilin isoforms (PS1 and PS2) containing γ-secretase show contrasting sensitivity to MRK-560. Here, the authors determined cryo-EM structures of PS1/PS2-complexes treated with MRK-560 and identified key residues responsible for the isoform-dependent selectivity.
- Xuefei Guo
- , Yumeng Wang
- & Yigong Shi
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Article
| Open AccessSpheroplasts preparation boosts the catalytic potential of a squalene-hopene cyclase
Squalene-hopene cyclase are membrane-bound enzymes used to produce bioactive compounds at industrial scale. Here, the authors show squalene-hopene cyclase spheroplasts, obtained by removing the outer cell membrane, having a higher catalytic activity than the enzymes and the whole cells, and their cross-linking for recycling and reuse.
- Ana I. Benítez-Mateos
- , Andreas Schneider
- & Francesca Paradisi
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Article
| Open AccessCryoEM structural exploration of catalytically active enzyme pyruvate carboxylase
Pyruvate Carboxylase is a multifunctional enzyme that follows a multi-pathway reaction. Here, the authors, using cryoEM and classification techniques, reveal several catalytic states at reaction sites, the interplay between them, and their relationship with motions in the tetrameric organization.
- Jorge Pedro López-Alonso
- , Melisa Lázaro
- & Mikel Valle
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Article
| Open AccessHelical ultrastructure of the metalloprotease meprin α in complex with a small molecule inhibitor
Meprin α is a proteolytic regulator of the extracellular matrix that forms enormous oligomeric filaments of unknown purpose. Here, the authors determine by cryo-EM the structural basis of the meprin supercoiled filament and further characterise a small molecule inhibitor bound to its active site.
- Charles Bayly-Jones
- , Christopher J. Lupton
- & James C. Whisstock
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Article
| Open AccessMammalian N1-adenosine PARylation is a reversible DNA modification
Poly-ADP-ribosylation (PARylation) is a well-known posttranslational modification of proteins. Here the authors show that beyond proteins also mammalian single-stranded DNA is PARylated in vitro and in vivo.
- Michael U. Musheev
- , Lars Schomacher
- & Christof Niehrs
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Article
| Open AccessStructure of the core human NADPH oxidase NOX2
NADPH oxidase NOX2 produces superoxide, a reactive oxygen species essential in innate immunity. Here, the authors reveal the structure of the NOX2 core, rationalize disease-causing mutations, and suggest avenues for selective NOX2 pharmacological modulation.
- Sigrid Noreng
- , Naruhisa Ota
- & James T. Koerber
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Article
| Open AccessWax worm saliva and the enzymes therein are the key to polyethylene degradation by Galleria mellonella
The crucial first step in the biodegradation of polyethylene plastic is oxidation of the polymer. This has traditionally required abiotic pre-treatment, but now Bertocchini and colleagues report two wax worm enzymes capable of catalyzing this oxidation and subsequent degradation at room temperature.
- A. Sanluis-Verdes
- , P. Colomer-Vidal
- & F. Bertocchini
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Article
| Open AccessMultiscale profiling of protease activity in cancer
The activity of multiple enzymes is dysregulated in cancer, but this cannot always be measured through enzyme expression. Here, the authors develop methods to measure protease activity across the organism, tissue, and single cell scales, and identify protease dysregulation in lung cancer and its response to targeted therapy.
- Ava P. Amini
- , Jesse D. Kirkpatrick
- & Sangeeta N. Bhatia
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Article
| Open AccessPhosphorylation of Jhd2 by the Ras-cAMP-PKA(Tpk2) pathway regulates histone modifications and autophagy
How cells transduce nutrient availability to appropriate gene expression remains poorly understood. Here the authors show that the nutrient sensor, protein kinase A modulates histone modifications and gene transcription by phosphorylating histone demethylase.
- Qi Yu
- , Xuanyunjing Gong
- & Shanshan Li
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Article
| Open AccessStructural characterization of a soil viral auxiliary metabolic gene product – a functional chitosanase
Metagenomics is revealing auxiliary metabolic genes (AMGs) in soil viral genomes. Here, authors solve the crystal structure for a soil viral AMG product, free and ligand bound, and show the protein can decompose chitin, a common carbon polymer.
- Ruonan Wu
- , Clyde A. Smith
- & Janet K. Jansson
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Article
| Open AccessStructural insights into the pSer/pThr dependent regulation of the SHP2 tyrosine phosphatase in insulin and CD28 signaling
SHP2 is an important human tyrosine phosphatase with key roles in cancer, immune responses and insulin signaling. Here, the authors explore its substrate recognition mechanism in molecular detail and uncover a complex regulatory mechanism for this enzyme that marks specific target sites for dephosphorylation.
- András Zeke
- , Tamás Takács
- & Attila Reményi
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Article
| Open AccessBiochemical and structural insights of multifunctional flavin-dependent monooxygenase FlsO1-catalyzed unexpected xanthone formation
The biosynthesis of xanthones has not been well documented. Here, the authors report that monooxygenase FlsO1 catalyzes three successive oxidations – hydroxylation, epoxidation and Baeyer–Villiger oxidation—to form the xanthone scaffold in actinomycetes.
- Chunfang Yang
- , Liping Zhang
- & Changsheng Zhang
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Article
| Open AccessStructural basis for APE1 processing DNA damage in the nucleosome
AP endonuclease 1 (APE1) processes genomic AP sites during base excision repair. Here, the authors determine the structural mechanism used by APE1 to process nucleosomal AP sites, providing new insight into DNA repair in chromatin.
- Tyler M. Weaver
- , Nicole M. Hoitsma
- & Bret D. Freudenthal
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Article
| Open AccessElucidation of divergent desaturation pathways in the formation of vinyl isonitrile and isocyanoacrylate
While canonical oxygen-rebound pathway has been utilized by several Fe/2OG enzymes, insights into the desaturation process remain to be elucidated. Here, the authors suggest that a carbocation might be deployed to enable chemically divergent desaturations through decarboxylation and deprotonation.
- Wantae Kim
- , Tzu-Yu Chen
- & Wei-chen Chang
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Article
| Open AccessEnzymes and cellular interplay required for flux of fixed nitrogen to ureides in bean nodules
Tropical legumes export fixed nitrogen from nodules as ureides. Here, the authors describe how ureides are produced by several biosynthetic enzymes in different nodule cell types and provide explanations for metabolic compartmentation.
- Luisa Voß
- , Katharina J. Heinemann
- & Claus-Peter Witte
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Article
| Open AccessSubstrate multiplexed protein engineering facilitates promiscuous biocatalytic synthesis
Efficient engineering of enzymes for expanded substrate scope is currently challenging. Here, the authors develop simple principles of how to design and interpret Substrate Multiplexed Screening assays to guide protein engineering to enable activity improvements with simultaneously with multiple substrates.
- Allwin D. McDonald
- , Peyton M. Higgins
- & Andrew R. Buller
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Article
| Open AccessStructure and activity of particulate methane monooxygenase arrays in methanotrophs
Particulate methane monooxygenase (pMMO) is the main enzyme used by methanotrophs. Here, the authors determined the native structure of pMMO by cryo-electron tomography, revealing lipid-stabilized features and a higher-order hexagonal array arrangement in intact cells.
- Yanan Zhu
- , Christopher W. Koo
- & Peijun Zhang