I03-Macromolecular Crystallography
I04-Macromolecular Crystallography
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Eugene
Kuatsjah
,
Alexa
Schwartz
,
Michael
Zahn
,
Konstantinos
Tornesakis
,
Zoe A.
Kellermyer
,
Morgan A.
Ingraham
,
Sean P.
Woodworth
,
Kelsey J.
Ramirez
,
Paul A.
Cox
,
Andrew R.
Pickford
,
Davinia
Salvachúa
Diamond Proposal Number(s):
[23269]
Open Access
Abstract: White-rot fungi (WRF) are the most efficient lignin-degrading organisms in nature. However, their capacity to use lignin-related aromatic compounds, such as 4-hydroxybenzoate, as carbon sources has only been described recently. Previously, the hydroxyquinol pathway was proposed for the bioconversion of these compounds in fungi, but gene- and structure-function relationships of the full enzymatic pathway remain uncharacterized in any single fungal species. Here, we characterize seven enzymes from two WRF, Trametes versicolor and Gelatoporia subvermispora, which constitute a four-enzyme cascade from 4-hydroxybenzoate to β-ketoadipate via the hydroxyquinol pathway. Furthermore, we solve the crystal structure of four of these enzymes and identify mechanistic differences with the closest bacterial and fungal structural homologs. Overall, this research expands our understanding of aromatic catabolism by WRF and establishes an alternative strategy for the conversion of lignin-related compounds to the valuable molecule β-ketoadipate, contributing to the development of biological processes for lignin valorization.
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Dec 2024
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I11-High Resolution Powder Diffraction
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Diamond Proposal Number(s):
[25166]
Open Access
Abstract: Solid-state electrolytes (SEs) hold the potential to overcome challenges that hinder the commercialization of lithium-sulfur batteries. However, their limited ionic conductivity makes lithium (Li) ion transport a critical bottleneck in accessing the superior capacity of sulfur. In this study, we investigate high-entropy Cl,Br-co-doped sulfide argyrodites as SEs in Li-sulfur (S) all-solid-state batteries (ASSBs).
exhibits an ionic conductivity of 6.9 mS
at room temperature with an activation energy of 0.28 eV, thanks to its high configurational entropy. The electrolyte with suitable cathodic stability and high ionic conductivity allows S composite cathodes to deliver an areal capacity over 6 mAh
when cycled with Li-In anodes. The reduced porosity of the SE separator and the formation of a passivating interphase with metallic Li enable Li-S ASSBs to cycle without short circuits, achieving an S utilization of over 75% at 0.1 C.
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Oct 2024
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I03-Macromolecular Crystallography
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Sarah
Hulin-Curtis
,
James K.
Geary
,
Bruce J.
Maclachlan
,
Danny M.
Altmann
,
Laury
Baillon
,
David K.
Cole
,
Alexander
Greenshields-Watson
,
Sophie J.
Hesketh
,
Ian R.
Humphreys
,
Ian M.
Jones
,
Sarah N.
Lauder
,
Georgina H.
Mason
,
Kathryn
Smart
,
D. Oliver
Scourfield
,
Jake
Scott
,
Ksenia
Sukhova
,
Richard J.
Stanton
,
Aaron
Wall
,
Pierre J.
Rizkallah
,
Wendy S.
Barclay
,
Awen
Gallimore
,
Andrew
Godkin
Diamond Proposal Number(s):
[10462]
Open Access
Abstract: CD4+ T cells are central to adaptive immunity. Their role in cross-protection in viral infections such as influenza and severe acute respiratory syndrome (SARS) is well documented; however, molecular rules governing T cell receptor (TCR) engagement of peptide-human leukocyte antigen (pHLA) class II are less understood. Here, we exploit an aspect of HLA class II presentation, the peptide-flanking residues (PFRs), to “tune” CD4+ T cell responses within an in vivo model system of influenza. Using a recombinant virus containing targeted substitutions at immunodominant HLA-DR1 epitopes, we demonstrate limited weight loss and improved clinical scores after heterosubtypic re-challenge. We observe enhanced protection linked to lung-derived influenza-specific CD4+ and CD8+ T cells prior to re-infection. Structural analysis of the ternary TCR:pHLA complex identifies that flanking amino acids influence side chains in the core 9-mer peptide, increasing TCR affinity. Augmentation of CD4+ T cell immunity is achievable with a single mutation, representing a strategy to enhance adaptive immunity that is decoupled from vaccine modality.
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Jun 2024
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I03-Macromolecular Crystallography
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Chang
Liu
,
Daming
Zhou
,
Aiste
Dijokaite-Guraliuc
,
Piyada
Supasa
,
Helen M. E.
Duyvesteyn
,
Helen M.
Ginn
,
Muneeswaran
Selvaraj
,
Alexander J.
Mentzer
,
Raksha
Das
,
Thushan I.
De Silva
,
Thomas G.
Ritter
,
Megan
Plowright
,
Thomas A.h.
Newman
,
Lizzie
Stafford
,
Barbara
Kronsteiner
,
Nigel
Temperton
,
Yuan
Lui
,
Martin
Fellermeyer
,
Philip
Goulder
,
Paul
Klenerman
,
Susanna J.
Dunachie
,
Michael I.
Barton
,
Mikhail A.
Kutuzov
,
Omer
Dushek
,
Elizabeth E.
Fry
,
Juthathip
Mongkolsapaya
,
Jingshan
Ren
,
David I.
Stuart
,
Gavin R.
Screaton
Diamond Proposal Number(s):
[28534, 27009]
Open Access
Abstract: BA.2.86, a recently described sublineage of SARS-CoV-2 Omicron, contains many mutations in the spike gene. It appears to have originated from BA.2 and is distinct from the XBB variants responsible for many infections in 2023. The global spread and plethora of mutations in BA.2.86 has caused concern that it may possess greater immune-evasive potential, leading to a new wave of infection. Here, we examine the ability of BA.2.86 to evade the antibody response to infection using a panel of vaccinated or naturally infected sera and find that it shows marginally less immune evasion than XBB.1.5. We locate BA.2.86 in the antigenic landscape of recent variants and look at its ability to escape panels of potent monoclonal antibodies generated against contemporary SARS-CoV-2 infections. We demonstrate, and provide a structural explanation for, increased affinity of BA.2.86 to ACE2, which may increase transmissibility.
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May 2024
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I12-JEEP: Joint Engineering, Environmental and Processing
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Diamond Proposal Number(s):
[29851]
Open Access
Abstract: The formation of heterogeneous Li structures at the anode/solid polymer electrolyte (SPE) membrane interphase of solid-state Li-metal batteries (SSLMBs) is one of the key factors that impede SSLMB performance. The relationship between Li+-ion transport kinetics and Li0 structural evolution at the buried interphase is critical but challenging to characterize. Here, we report an operando correlative X-ray Compton scattering and computed tomography imaging technique that quantifies the changes of Li+-ion concentrations in the bulk cathode, SPE membrane, and anode of the SSLMB full cell using a commercially standard configuration. We then visualize Li+-ion concentration distributions as well as Li0 microstructures at the buried anode/SPE interphase. Mechanistic analyses show that the Li-stripping step forms more irregular interfacial Li morphologies at the expense of bulk anode volume shrinkage compared to the Li-plating step during the first cycle.
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May 2024
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B23-Circular Dichroism
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Diamond Proposal Number(s):
[31640]
Open Access
Abstract: The accumulation of plastic waste in the environment is an ecological disaster that requires a plurality of approaches to tackle. There is therefore an ever-pressing need to close the loop on production of both conventional and bioderived plastics. In response, we propose a generalizable biocatalysis engineering strategy to enhance the use of enzymes to depolymerize a broad class of plastics. To demonstrate this approach, we have solubilized and stabilized the commonly available lipase B from Candida antarctica in ionic liquids. In doing so, we unlock the ability of the enzyme to hydrolytically depolymerize post-consumer poly(lactic acid) plastic, demonstrating full degradation within 24 h and full conversion to monomer within 40 h at 90°C. This facile and scalable modification strategy allows for elevated operation temperatures in combination with the superior solvent capabilities of ionic liquids, providing a blueprint for enhancing the capability of any hydrolytic enzyme for plastic recycling.
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Jan 2024
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I04-1-Macromolecular Crystallography (fixed wavelength)
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Ting-Yu
Lin
,
Shakti
Ramsamooj
,
Tiffany
Perrier
,
Katarina
Liberatore
,
Louise
Lantier
,
Neil
Vasan
,
Kannan
Karukurichi
,
Seo-Kyoung
Hwang
,
Edward A.
Kesicki
,
Edward R.
Kastenhuber
,
Thorsten
Wiederhold
,
Tomer M.
Yaron
,
Emily M.
Huntsman
,
Mengmeng
Zhu
,
Yilun
Ma
,
Marcia N.
Paddock
,
Guoan
Zhang
,
Benjamin D.
Hopkins
,
Owen
Mcguinness
,
Robert E.
Schwartz
,
Baran A.
Ersoy
,
Lewis C.
Cantley
,
Jared L.
Johnson
,
Marcus D.
Goncalves
Open Access
Abstract: The phosphoinositide 3-kinase p110α is an essential mediator of insulin signaling and glucose homeostasis. We interrogated the human serine, threonine, and tyrosine kinome to search for novel regulators of p110α and found that the Hippo kinases phosphorylate p110α at T1061, which inhibits its activity. This inhibitory state corresponds to a conformational change of a membrane-binding domain on p110α, which impairs its ability to engage membranes. In human primary hepatocytes, cancer cell lines, and rodent tissues, activation of the Hippo kinases MST1/2 using forskolin or epinephrine is associated with phosphorylation of T1061 and inhibition of p110α, impairment of downstream insulin signaling, and suppression of glycolysis and glycogen synthesis. These changes are abrogated when MST1/2 are genetically deleted or inhibited with small molecules or if the T1061 is mutated to alanine. Our study defines an inhibitory pathway of PI3K signaling and a link between epinephrine and insulin signaling.
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Dec 2023
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I04-1-Macromolecular Crystallography (fixed wavelength)
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Heather L.
Martin
,
Amy L.
Turner
,
Julie
Higgins
,
Anna A.
Tang
,
Christian
Tiede
,
Thomas
Taylor
,
Sitthinon
Siripanthong
,
Thomas L.
Adams
,
Iain W.
Manfield
,
Sandra M.
Bell
,
Ewan E.
Morrison
,
Jacquelyn
Bond
,
Chi H.
Trinh
,
Carolyn D.
Hurst
,
Margaret A.
Knowles
,
Richard W.
Bayliss
,
Darren C.
Tomlinson
Diamond Proposal Number(s):
[29704]
Open Access
Abstract: Kinases are important therapeutic targets, and their inhibitors are classified according to their mechanism of action, which range from blocking ATP binding to covalent inhibition. Here, a mechanism of inhibition is highlighted by capturing p21-activated kinase 5 (PAK5) in an intermediate state of activation using an Affimer reagent that binds in the P+1 pocket. PAK5 was identified from a non-hypothesis-driven high-content imaging RNAi screen in urothelial cancer cells. Silencing of PAK5 resulted in reduced cell number, G1/S arrest, and enlargement of cells, suggesting it to be important in urothelial cancer cell line survival and proliferation. Affimer reagents were isolated to identify mechanisms of inhibition. The Affimer PAK5-Af17 recapitulated the phenotype seen with siRNA. Co-crystallization revealed that PAK5-Af17 bound in the P+1 pocket of PAK5, locking the kinase into a partial activation state. This mechanism of inhibition indicates that another class of kinase inhibitors is possible.
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Oct 2023
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I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
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Yuan
Chen
,
Georgina H.
Mason
,
D. Oliver
Scourfield
,
Alexander
Greenshields-Watson
,
Tracey A.
Haigh
,
Andrew K.
Sewell
,
Heather M.
Long
,
Awen M.
Gallimore
,
Pierre
Rizkallah
,
Bruce J.
Maclachlan
,
Andrew
Godkin
Diamond Proposal Number(s):
[20147, 29502, 29990]
Open Access
Abstract: CD4+ T cells recognize a broad range of peptide epitopes of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which contribute to immune memory and limit COVID-19 disease. We demonstrate that the immunogenicity of SARS-CoV-2 peptides, in the context of the model allotype HLA-DR1, does not correlate with their binding affinity to the HLA heterodimer. Analyzing six epitopes, some with very low binding affinity, we solve X-ray crystallographic structures of each bound to HLA-DR1. Further structural definitions reveal the precise molecular impact of viral variant mutations on epitope presentation. Omicron escaped ancestral SARS-CoV-2 immunity to two epitopes through two distinct mechanisms: (1) mutations to TCR-facing epitope positions and (2) a mechanism whereby a single amino acid substitution caused a register shift within the HLA binding groove, completely altering the peptide-HLA structure. This HLA-II-specific paradigm of immune escape highlights how CD4+ T cell memory is finely poised at the level of peptide-HLA-II presentation.
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Aug 2023
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B21-High Throughput SAXS
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Open Access
Abstract: Bacteriophage T3 encodes a SAMase that, through cleavage of S-adenosyl methionine (SAM), circumvents the SAM-dependent type I restriction-modification (R-M) defense. We show that SAMase also allows T3 to evade the BREX defense. Although SAM depletion weakly affects BREX methylation, it completely inhibits the defensive function of BREX, suggesting that SAM could be a co-factor for BREX-mediated exclusion of phage DNA, similar to its anti-defense role in type I R-M. The anti-BREX activity of T3 SAMase is mediated not just by enzymatic degradation of SAM but also by direct inhibition of MetK, the host SAM synthase. We present a 2.8 Å cryoelectron microscopy (cryo-EM) structure of the eight-subunit T3 SAMase-MetK complex. Structure-guided mutagenesis reveals that this interaction stabilizes T3 SAMase in vivo, further stimulating its anti-BREX activity. This work provides insights in the versatility of bacteriophage counterdefense mechanisms and highlights the role of SAM as a co-factor of diverse bacterial immunity systems.
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Aug 2023
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