Search Results

You searched for all publications:

with publication states 'Published (Approved)'
published in journal 'Life'

Search Again...

These results only include publications that have been reviewed and processed by Diamond Light Source. To also view papers that have not yet been processed click here.

You can use the folder icon under Actions to collate papers as required. You can then click on View Folder in the left-hand navigation to review and/or download your folder.

Exact matches
Related results

21 items found (0 items not approved), displaying 1 to 10.[First/Prev] 1, 2, 3 [Next/Last]

Instruments / Technical Area	Publication Details	Published
I03-Macromolecular Crystallography	H2S biogenesis by cystathionine beta-synthase: mechanism of inhibition by aminooxyacetic acid and unexpected role of serine Maria Petrosino , Karim Zuhra , Jola Kopec , Andrew Hutchin , Csaba Szabo , Tomas Majtan Cellular And Molecular Life Sciences 79 DOI: 10.1007/s00018-022-04479-9 Diamond Proposal Number(s): [29346] Open Access Show Abstract ▼ Abstract: Cystathionine beta-synthase (CBS) is a pivotal enzyme of the transsulfuration pathway responsible for diverting homocysteine to the biosynthesis of cysteine and production of hydrogen sulfide (H2S). Aberrant upregulation of CBS and overproduction of H2S contribute to pathophysiology of several diseases including cancer and Down syndrome. Therefore, pharmacological CBS inhibition has emerged as a prospective therapeutic approach. Here, we characterized binding and inhibitory mechanism of aminooxyacetic acid (AOAA), the most commonly used CBS inhibitor. We found that AOAA binds CBS tighter than its respective substrates and forms a dead-end PLP-bound intermediate featuring an oxime bond. Surprisingly, serine, but not cysteine, replaced AOAA from CBS and formed an aminoacrylate reaction intermediate, which allowed for the continuation of the catalytic cycle. Indeed, serine rescued and essentially normalized the enzymatic activity of AOAA-inhibited CBS. Cellular studies confirmed that AOAA decreased H2S production and bioenergetics, while additional serine rescued CBS activity, H2S production and mitochondrial function. The crystal structure of AOAA-bound human CBS showed a lack of hydrogen bonding with residues G305 and Y308, found in the serine-bound model. Thus, AOAA-inhibited CBS could be reactivated by serine. This difference may be important in a cellular environment in multiple pathophysiological conditions and may modulate the CBS-inhibitory activity of AOAA. In addition, our results demonstrate additional complexities of using AOAA as a CBS-specific inhibitor of H2S biogenesis and point to the urgent need to develop a potent, selective and specific pharmacological CBS inhibitor.	Aug 2022
B21-High Throughput SAXS I04-1-Macromolecular Crystallography (fixed wavelength)	Structure and function of an atypical homodimeric actin capping protein from the malaria parasite Abris Adam Bendes , Petri Kursula , Inari Kursula Cellular And Molecular Life Sciences 79 DOI: 10.1007/s00018-021-04032-0 Open Access Show Abstract ▼ Abstract: Apicomplexan parasites, such as Plasmodium spp., rely on an unusual actomyosin motor, termed glideosome, for motility and host cell invasion. The actin filaments are maintained by a small set of essential regulators, which provide control over actin dynamics in the different stages of the parasite life cycle. Actin filament capping proteins (CPs) are indispensable heterodimeric regulators of actin dynamics. CPs have been extensively characterized in higher eukaryotes, but their role and functional mechanism in Apicomplexa remain enigmatic. Here, we present the first crystal structure of a homodimeric CP from the malaria parasite and compare the homo- and heterodimeric CP structures in detail. Despite retaining several characteristics of a canonical CP, the homodimeric Plasmodium berghei (Pb)CP exhibits crucial differences to the canonical heterodimers. Both homo- and heterodimeric PbCPs regulate actin dynamics in an atypical manner, facilitating rapid turnover of parasite actin, without affecting its critical concentration. Homo- and heterodimeric PbCPs show partially redundant activities, possibly to rescue actin filament capping in life cycle stages where the β-subunit is downregulated. Our data suggest that the homodimeric PbCP also influences actin kinetics by recruiting lateral actin dimers. This unusual function could arise from the absence of a β-subunit, as the asymmetric PbCP homodimer lacks structural elements essential for canonical barbed end interactions suggesting a novel CP binding mode. These findings will facilitate further studies aimed at elucidating the precise actin filament capping mechanism in Plasmodium.	Feb 2022
I03-Macromolecular Crystallography	PSTPIP1-LYP phosphatase interaction: structural basis and implications for autoinflammatory disorders Jose Antonio Manso , Tamara Marcos , Virginia Ruiz-Martín , Javier Casas , Pablo Alcon , Mariano Sánchez Crespo , Yolanda Bayón , Jose M. De Pereda , Andrés Alonso Cellular And Molecular Life Sciences 79 DOI: 10.1007/s00018-022-04173-w Diamond Proposal Number(s): [10121] Open Access Show Abstract ▼ Abstract: Mutations in the adaptor protein PSTPIP1 cause a spectrum of autoinflammatory diseases, including PAPA and PAMI; however, the mechanism underlying these diseases remains unknown. Most of these mutations lie in PSTPIP1 F-BAR domain, which binds to LYP, a protein tyrosine phosphatase associated with arthritis and lupus. To shed light on the mechanism by which these mutations generate autoinflammatory disorders, we solved the structure of the F-BAR domain of PSTPIP1 alone and bound to the C-terminal homology segment of LYP, revealing a novel mechanism of recognition of Pro-rich motifs by proteins in which a single LYP molecule binds to the PSTPIP1 F-BAR dimer. The residues R228, D246, E250, and E257 of PSTPIP1 that are mutated in immunological diseases directly interact with LYP. These findings link the disruption of the PSTPIP1/LYP interaction to these diseases, and support a critical role for LYP phosphatase in their pathogenesis.	Feb 2022
	The high-throughput production of membrane proteins James Birch , Andrew Quigley Emerging Topics In Life Sciences 6 DOI: 10.1042/ETLS20210196 Open Access Show Abstract ▼ Abstract: Membrane proteins, found at the junctions between the outside world and the inner workings of the cell, play important roles in human disease and are used as biosensors. More than half of all therapeutics directly affect membrane protein function while nanopores enable DNA sequencing. The structural and functional characterisation of membrane proteins is therefore crucial. However, low levels of naturally abundant protein and the hydrophobic nature of membrane proteins makes production difficult. To maximise success, high-throughput strategies were developed that rely upon simple screens to identify successful constructs and rapidly exclude those unlikely to work. Parameters that affect production such as expression host, membrane protein origin, expression vector, fusion-tags, encapsulation reagent and solvent composition are screened in parallel. In this way, constructs with divergent requirements can be produced for a variety of structural applications. As structural techniques advance, sample requirements will change. Single-particle cryo-electron microscopy requires less protein than crystallography and as cryo-electron tomography and time-resolved serial crystallography are developed new sample production requirements will evolve. Here we discuss different methods used for the high-throughput production of membrane proteins for structural biology.	Oct 2021
B21-High Throughput SAXS	The structure and flexibility analysis of the Arabidopsis synaptotagmin 1 reveal the basis of its regulation at membrane contact sites Juan L Benavente , Dritan Siliqi , Lourdes Infantes , Laura Lagartera , Alberto Mills , Federico Gago , Noemí Ruiz-López , Miguel A Botella , Maria J. Sanchez-Barrena , Armando Albert Life Science Alliance 4 DOI: 10.26508/lsa.202101152 Diamond Proposal Number(s): [21741] Open Access Show Abstract ▼ Abstract: Non-vesicular lipid transfer at ER and plasma membrane (PM) contact sites (CS) is crucial for the maintenance of membrane lipid homeostasis. Extended synaptotagmins (E-Syts) play a central role in this process as they act as molecular tethers of ER and PM and as lipid transfer proteins between these organelles. E-Syts are proteins constitutively anchored to the ER through an N-terminal hydrophobic segment and bind the PM via a variable number of C-terminal C2 domains. Synaptotagmins (SYTs) are the plant orthologous of E-Syts and regulate the ER–PM communica- tion in response to abiotic stress. Combining different structural and biochemical techniques, we demonstrate that the binding of SYT1 to lipids occurs through a Ca2+-dependent lipid-binding site and by a site for phosphorylated forms of phosphatidylinositol, thus integrating two different molecular signals in response to stress. In addition, we show that SYT1 displays three highly flexible hinge points that provide conformational freedom to facilitate lipid extraction, protein loading, and subsequent transfer between PM and ER.	Aug 2021
B24-Cryo Soft X-ray Tomography	Single cell cryo-soft X-ray tomography shows that each Chlamydia trachomatis inclusion is a unique community of bacteria Patrick Phillips , James M. Parkhurst , Ilias Kounatidis , Chidinma Okolo , Thomas M. Fish , James H. Naismith , Martin A. Walsh , Maria Harkiolaki , Maud Dumoux Life 11 DOI: 10.3390/life11080842 Open Access Show Abstract ▼ Abstract: Chlamydiae are strict intracellular pathogens residing within a specialised membrane-bound compartment called the inclusion. Therefore, each infected cell can, be considered as a single entity where bacteria form a community within the inclusion. It remains unclear as to how the population of bacteria within the inclusion influences individual bacterium. The life cycle of Chlamydia involves transitioning between the invasive elementary bodies (EBs) and replicative reticulate bodies (RBs). We have used cryo-soft X-ray tomography to observe individual inclusions, an approach that combines 40 nm spatial resolution and large volume imaging (up to 16 µm). Using semi-automated segmentation pipeline, we considered each inclusion as an individual bacterial niche. Within each inclusion, we identifyed and classified different forms of the bacteria and confirmed the recent finding that RBs have a variety of volumes (small, large and abnormal). We demonstrate that the proportions of these different RB forms depend on the bacterial concentration in the inclusion. We conclude that each inclusion operates as an autonomous community that influences the characteristics of individual bacteria within the inclusion.	Aug 2021
I04-1-Macromolecular Crystallography (fixed wavelength) I04-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	Structural comparison of GLUT1 to GLUT3 reveal transport regulation mechanism in sugar porter family Tania Filipa Custodio , Peter Aasted Paulsen , Kelly May Frain , Bjorn Panyella Pedersen Life Science Alliance 4 DOI: 10.26508/lsa.202000858 Diamond Proposal Number(s): [23316, 17844, 13062] Show Abstract ▼ Abstract: The human glucose transporters GLUT1 and GLUT3 have a central role in glucose uptake as canonical members of the Sugar Porter (SP) family. GLUT1 and GLUT3 share a fully conserved substrate-binding site with identical substrate coordination, but differ significantly in transport affinity in line with their physiological function. Here, we present a 2.4 Å crystal structure of GLUT1 in an inward open conformation and compare it with GLUT3 using both structural and functional data. Our work shows that interactions between a cytosolic “SP motif” and a conserved “A motif” stabilize the outward conformational state and increases substrate apparent affinity. Furthermore, we identify a previously undescribed Cl− ion site in GLUT1 and an endofacial lipid/glucose binding site which modulate GLUT kinetics. The results provide a possible explanation for the difference between GLUT1 and GLUT3 glucose affinity, imply a general model for the kinetic regulation in GLUTs and suggest a physiological function for the defining SP sequence motif in the SP family.	Feb 2021
I03-Macromolecular Crystallography	Structure of the helicase core of Werner helicase, a key target in microsatellite instability cancers Joseph A Newman , Angeline E. Gavard , Simone Lieb , Madhwesh C. Ravichandran , Katja Hauer , Patrick Werni , Leonhard Geist , Jark Böttcher , John R Engen , Klaus Rumpel , Matthias Samwer , Mark Petronczki , Opher Gileadi Life Science Alliance 4 DOI: 10.26508/lsa.202000795 Diamond Proposal Number(s): [19301] Open Access Show Abstract ▼ Abstract: Loss of WRN, a DNA repair helicase, was identified as a strong vulnerability of microsatellite instable (MSI) cancers, making WRN a promising drug target. We show that ATP binding and hydrolysis are required for genome integrity and viability of MSI cancer cells. We report a 2.2-Å crystal structure of the WRN helicase core (517–1,093), comprising the two helicase subdomains and winged helix domain but not the HRDC domain or nuclease domains. The structure highlights unusual features. First, an atypical mode of nucleotide binding that results in unusual relative positioning of the two helicase subdomains. Second, an additional β-hairpin in the second helicase subdomain and an unusual helical hairpin in the Zn2+ binding domain. Modelling of the WRN helicase in complex with DNA suggests roles for these features in the binding of alternative DNA structures. NMR analysis shows a weak interaction between the HRDC domain and the helicase core, indicating a possible biological role for this association. Together, this study will facilitate the structure-based development of inhibitors against WRN helicase.	Nov 2020
I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength)	Implications for tetraspanin-enriched microdomain assembly based on structures of CD9 with EWI-F Wout Oosterheert , Katerina T. Xenaki , Viviana Neviani , Wouter Pos , Sofia Doulkeridou , Jip Manshande , Nicholas M. Pearce , Loes M. J. Kroon-Batenburg , Martin Lutz , Paul M. P. Van Bergen En Henegouwen , Piet Gros Life Science Alliance 3 DOI: 10.26508/lsa.202000883 Diamond Proposal Number(s): [19800] Open Access Show Abstract ▼ Abstract: Tetraspanins are eukaryotic membrane proteins that contribute to a variety of signaling processes by organizing partner-receptor molecules in the plasma membrane. How tetraspanins bind and cluster partner receptors into tetraspanin-enriched microdomains is unknown. Here, we present crystal structures of the large extracellular loop of CD9 bound to nanobodies 4C8 and 4E8 and, the cryo-EM structure of 4C8-bound CD9 in complex with its partner EWI-F. CD9–EWI-F displays a tetrameric arrangement with two central EWI-F molecules, dimerized through their ectodomains, and two CD9 molecules, one bound to each EWI-F transmembrane helix through CD9-helices h3 and h4. In the crystal structures, nanobodies 4C8 and 4E8 bind CD9 at loops C and D, which is in agreement with the 4C8 conformation in the CD9–EWI-F complex. The complex varies from nearly twofold symmetric (with the two CD9 copies nearly anti-parallel) to ca. 50° bent arrangements. This flexible arrangement of CD9–EWI-F with potential CD9 homo-dimerization at either end provides a “concatenation model” for forming short linear or circular assemblies, which may explain the occurrence of tetraspanin-enriched microdomains.	Sep 2020
I04-1-Macromolecular Crystallography (fixed wavelength) I24-Microfocus Macromolecular Crystallography	STAG1 vulnerabilities for exploiting cohesin synthetic lethality in STAG2-deficient cancers Petra Van Der Lelij , Joseph A. Newman , Simone Lieb , Julian Jude , Vittorio Katis , Thomas Hoffmann , Matthias Hinterndorfer , Gerd Bader , Norbert Kraut , Mark A. Pearson , Jan-Michael Peters , Johannes Zuber , Opher Gileadi , Mark Petronczki Life Science Alliance 3 DOI: 10.26508/lsa.202000725 Diamond Proposal Number(s): [19301, 18145] Open Access Show Abstract ▼ Abstract: The cohesin subunit STAG2 has emerged as a recurrently inactivated tumor suppressor in human cancers. Using candidate approaches, recent studies have revealed a synthetic lethal interaction between STAG2 and its paralog STAG1. To systematically probe genetic vulnerabilities in the absence of STAG2, we have performed genome-wide CRISPR screens in isogenic cell lines and identified STAG1 as the most prominent and selective dependency of STAG2-deficient cells. Using an inducible degron system, we show that chemical genetic degradation of STAG1 protein results in the loss of sister chromatid cohesion and rapid cell death in STAG2-deficient cells, while sparing STAG2–wild-type cells. Biochemical assays and X-ray crystallography identify STAG1 regions that interact with the RAD21 subunit of the cohesin complex. STAG1 mutations that abrogate this interaction selectively compromise the viability of STAG2-deficient cells. Our work highlights the degradation of STAG1 and inhibition of its interaction with RAD21 as promising therapeutic strategies. These findings lay the groundwork for the development of STAG1-directed small molecules to exploit synthetic lethality in STAG2-mutated tumors.	May 2020

Publications Database

Search Results

You searched for all publications:

Search Again...

Subject Areas (check all that apply) select all

Instruments used to collect data (check all that apply) select all

Collaborations involved (check all that apply) select all

Diamond Offline Facilities used

Relevant Technical Areas (check all that apply) select all

Menu for Anonymous User