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55 items found (0 items not approved), displaying 1 to 10.[First/Prev] 1, 2, 3, 4, 5, 6 [Next/Last]

Instruments / Technical Area	Publication Details	Published
I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength)	KorB switching from DNA-sliding clamp to repressor mediates long-range gene silencing in a multi-drug resistance plasmid Thomas C. Mclean , Francisco Balaguer-Pérez , Joshua Chandanani , Christopher M. Thomas , Clara Aicart-Ramos , Sophia Burick , Paul Dominic B. Olinares , Giulia Gobbato , Julia E. A. Mundy , Brian T. Chait , David M. Lawson , Seth A. Darst , Elizabeth A. Campbell , Fernando Moreno-Herrero , Tung Le Nature Microbiology 20 DOI: 10.1038/s41564-024-01915-3 Diamond Proposal Number(s): [25108] Open Access Show Abstract ▼ Abstract: Examples of long-range gene regulation in bacteria are rare and generally thought to involve DNA looping. Here, using a combination of biophysical approaches including X-ray crystallography and single-molecule analysis for the KorB–KorA system in Escherichia coli, we show that long-range gene silencing on the plasmid RK2, a source of multi-drug resistance across diverse Gram-negative bacteria, is achieved cooperatively by a DNA-sliding clamp, KorB, and a clamp-locking protein, KorA. We show that KorB is a CTPase clamp that can entrap and slide along DNA to reach distal target promoters up to 1.5 kb away. We resolved the tripartite crystal structure of a KorB–KorA–DNA co-complex, revealing that KorA latches KorB into a closed clamp state. DNA-bound KorA thus stimulates repression by stalling KorB sliding at target promoters to occlude RNA polymerase holoenzymes. Together, our findings explain the mechanistic basis for KorB role switching from a DNA-sliding clamp to a co-repressor and provide an alternative mechanism for long-range regulation of gene expression in bacteria.	Jan 2025
I04-Macromolecular Crystallography	Structural aspects of Mycobacterium tuberculosis DNA gyrase targeted by novel bacterial topoisomerase inhibitors Maja Kokot , Martina Hrast Rambaher , Lipeng Feng , Lesley A Mitchenall , David M. Lawson , Anthony Maxwell , Tanya Parish , Nikola Minovski , Marko Anderluh Acs Medicinal Chemistry Letters DOI: 10.1021/acsmedchemlett.4c00447 Diamond Proposal Number(s): [25108] Open Access Show Abstract ▼ Abstract: In this Letter, we present a small series of novel bacterial topoisomerase inhibitors (NTBIs) that exhibit both potent inhibition of Mycobacterium tuberculosis DNA gyrase and potent antimycobacterial activity. The disclosed crystal structure of M. tuberculosis DNA gyrase in complex with DNA and compound 5 from this NBTI series reveals the binding mode of an NBTI in the GyrA binding pocket and confirms the presence and importance of halogen bonding for the excellent on-target potency. In addition, we have shown that compound 5 is a promising M. tuberculosis DNA gyrase inhibitor, with an IC50 for M. tuberculosis gyrase of 0.096 μM, and it has potent activity against M. tuberculosis, with an IC50 of 0.165 μM.	Nov 2024
I04-Macromolecular Crystallography	Exploring the interaction of N-(benzothiazol-2-yl)pyrrolamide DNA gyrase inhibitors with the GyrB ATP-binding site lipophilic floor: A medicinal chemistry and QTAIM study Nace Zidar , Andrej Emanuel Cotman , Wessel Sinnige , Ondrej Benek , Michaela Barančokova , Anamarija Zega , Lucija Peterlin Mašič , Tihomir Tomašič , Janez Ilaš , Sara R. Henderson , Julia E. A. Mundy , Anthony Maxwell , Clare E. M. Stevenson , David M. Lawson , Geert Jan Sterk , Rodrigo Tosso , Lucas Gutierrez , Ricardo D. Enriz , Danijel Kikelj Bioorganic & Medicinal Chemistry 43 DOI: 10.1016/j.bmc.2024.117798 Diamond Proposal Number(s): [18565] Open Access Show Abstract ▼ Abstract: N-(Benzothiazole-2-yl)pyrrolamide DNA gyrase inhibitors with benzyl or phenethyl substituents attached to position 3 of the benzothiazole ring or to the carboxamide nitrogen atom were prepared and studied for their inhibition of Escherichia coli DNA gyrase by supercoiling assay. Compared to inhibitors bearing the substituents at position 4 of the benzothiazole ring, the inhibition was attenuated by moving the substituent to position 3 and further to the carboxamide nitrogen atom. A co-crystal structure of (Z)-3-benzyl-2-((4,5-dibromo-1H-pyrrole-2-carbonyl)imino)-2,3-dihydrobenzo[d]-thiazole-6-carboxylic acid (I) in complex with E. coli GyrB24 (ATPase subdomain) was solved, revealing the binding mode of this type of inhibitor to the ATP-binding pocket of the E. coli GyrB subunit. The key binding interactions were identified and their contribution to binding was rationalised by quantum theory of atoms in molecules (QTAIM) analysis. Our study shows that the benzyl or phenethyl substituents bound to the benzothiazole core interact with the lipophilic floor of the active site, which consists mainly of residues Gly101, Gly102, Lys103 and Ser108. Compounds with substituents at position 3 of the benzothiazole core were up to two orders of magnitude more effective than compounds with substituents at the carboxamide nitrogen. In addition, the 6-oxalylamino compounds were more potent inhibitors of E. coli DNA gyrase than the corresponding 6-acetamido analogues.	Jun 2024
I04-Macromolecular Crystallography	Bimodular architecture of bacterial effector SAP05 that drives ubiquitin-independent targeted protein degradation Qun Liu , Abbas Maqbool , Federico G. Mirkin , Yeshveer Singh , Clare E. M. Stevenson , David M. Lawson , Sophien Kamoun , Weijie Huang , Saskia A. Hogenhout Proceedings Of The National Academy Of Sciences 120 DOI: 10.1073/pnas.2310664120 Diamond Proposal Number(s): [25108] Open Access Show Abstract ▼ Abstract: In eukaryotes, targeted protein degradation (TPD) typically depends on a series of interactions among ubiquitin ligases that transfer ubiquitin molecules to substrates leading to degradation by the 26S proteasome. We previously identified that the bacterial effector protein SAP05 mediates ubiquitin-independent TPD. SAP05 forms a ternary complex via interactions with the von Willebrand Factor Type A (vWA) domain of the proteasomal ubiquitin receptor Rpn10 and the zinc-finger (ZnF) domains of the SQUAMOSA-PROMOTER BINDING PROTEIN-LIKE (SPL) and GATA BINDING FACTOR (GATA) transcription factors (TFs). This leads to direct TPD of the TFs by the 26S proteasome. Here, we report the crystal structures of the SAP05–Rpn10vWA complex at 2.17 Å resolution and of the SAP05–SPL5ZnF complex at 2.20 Å resolution. Structural analyses revealed that SAP05 displays a remarkable bimodular architecture with two distinct nonoverlapping surfaces, a “loop surface” with three protruding loops that form electrostatic interactions with ZnF, and a “sheet surface” featuring two β-sheets, loops, and α-helices that establish polar interactions with vWA. SAP05 binding to ZnF TFs involves single amino acids responsible for multiple contacts, while SAP05 binding to vWA is more stable due to the necessity of multiple mutations to break the interaction. In addition, positioning of the SAP05 complex on the 26S proteasome points to a mechanism of protein degradation. Collectively, our findings demonstrate how a small bacterial bimodular protein can bypass the canonical ubiquitin–proteasome proteolysis pathway, enabling ubiquitin-independent TPD in eukaryotic cells. This knowledge holds significant potential for the creation of TPD technologies.	Dec 2023
	The CCP 4 suite: integrative software for macromolecular crystallography Jon Agirre , Mihaela Atanasova , Haroldas Bagdonas , Charles B. Ballard , Arnaud Basle , James Beilsten-Edmands , Rafael J. Borges , David G. Brown , J. Javier Burgos-Marmol , John M. Berrisford , Paul S. Bond , Iracema Caballero , Lucrezia Catapano , Grzegorz Chojnowski , Atlanta G. Cook , Kevin D. Cowtan , Tristan I. Croll , Judit É. Debreczeni , Nicholas E. Devenish , Eleanor J. Dodson , Tarik R. Drevon , Paul Emsley , Gwyndaf Evans , Phil R. Evans , Maria Fando , James Foadi , Luis Fuentes-Montero , Elspeth F. Garman , Markus Gerstel , Richard J. Gildea , Kaushik Hatti , Maarten L. Hekkelman , Philipp Heuser , Soon Wen Hoh , Michael A. Hough , Huw T. Jenkins , Elisabet Jiménez , Robbie P. Joosten , Ronan M. Keegan , Nicholas Keep , Eugene B. Krissinel , Petr Kolenko , Oleg Kovalevskiy , Victor S. Lamzin , David M. Lawson , Andrey Lebedev , Andrew G. W. Leslie , Bernhard Lohkamp , Fei Long , Martin Maly , Airlie Mccoy , Stuart J. Mcnicholas , Ana Medina , Claudia Millán , James W. Murray , Garib N. Murshudov , Robert A. Nicholls , Martin E. M. Noble , Robert Oeffner , Navraj S. Pannu , James M. Parkhurst , Nicholas Pearce , Joana Pereira , Anastassis Perrakis , Harold R. Powell , Randy J. Read , Daniel J. Rigden , William Rochira , Massimo Sammito , Filomeno Sanchez Rodriguez , George M. Sheldrick , Kathryn L. Shelley , Felix Simkovic , Adam J. Simpkin , Pavol Skubak , Egor Sobolev , Roberto A. Steiner , Kyle Stevenson , Ivo Tews , Jens M. H. Thomas , Andrea Thorn , Josep Triviño Valls , Ville Uski , Isabel Uson , Alexei Vagin , Sameer Velankar , Melanie Vollmar , Helen Walden , David Waterman , Keith S. Wilson , Martyn Winn , Graeme Winter , Marcin Wojdyr , Keitaro Yamashita Acta Crystallographica Section D Structural Biology 79, 449 - 461 DOI: 10.1107/S2059798323003595 Open Access Show Abstract ▼ Abstract: The Collaborative Computational Project No. 4 (CCP4) is a UK-led international collective with a mission to develop, test, distribute and promote software for macromolecular crystallography. The CCP4 suite is a multiplatform collection of programs brought together by familiar execution routines, a set of common libraries and graphical interfaces. The CCP4 suite has experienced several considerable changes since its last reference article, involving new infrastructure, original programs and graphical interfaces. This article, which is intended as a general literature citation for the use of the CCP4 software suite in structure determination, will guide the reader through such transformations, offering a general overview of the new features and outlining future developments. As such, it aims to highlight the individual programs that comprise the suite and to provide the latest references to them for perusal by crystallographers around the world.	Jun 2023
I03-Macromolecular Crystallography	Exploring the 5-substituted 2-aminobenzothiazole-based DNA Gyrase B inhibitors active against ESKAPE pathogens Maša Sterle , Martina Durcik , Clare E. M. Stevenson , Sara Henderson , Petra Eva Szili , Marton Czikkely , David M. Lawson , Anthony Maxwell , Dominique Cahard , Danijel Kikelj , Nace Zidar , Csaba Pal , Lucija Peterlin Mašič , Janez Ilaš , Tihomir Tomašič , Andrej Emanuel Cotman , Anamarija Zega Acs Omega DOI: 10.1021/acsomega.3c01930 Diamond Proposal Number(s): [18565] Open Access Show Abstract ▼ Abstract: We present a new series of 2-aminobenzothiazole-based DNA gyrase B inhibitors with promising activity against ESKAPE bacterial pathogens. Based on the binding information extracted from the cocrystal structure of DNA gyrase B inhibitor A, in complex with Escherichia coli GyrB24, we expanded the chemical space of the benzothiazole-based series to the C5 position of the benzothiazole ring. In particular, compound E showed low nanomolar inhibition of DNA gyrase (IC50 < 10 nM) and broad-spectrum antibacterial activity against pathogens belonging to the ESKAPE group, with the minimum inhibitory concentration < 0.03 μg/mL for most Gram-positive strains and 4–16 μg/mL against Gram-negative E. coli, Acinetobacter baumannii, Pseudomonas aeruginosa, and Klebsiella pneumoniae. To understand the binding mode of the synthesized inhibitors, a combination of docking calculations, molecular dynamics (MD) simulations, and MD-derived structure-based pharmacophore modeling was performed. The computational analysis has revealed that the substitution at position C5 can be used to modify the physicochemical properties and antibacterial spectrum and enhance the inhibitory potency of the compounds. Additionally, a discussion of challenges associated with the synthesis of 5-substituted 2-aminobenzothiazoles is presented.	Jun 2023
I03-Macromolecular Crystallography	Resurrection of plant disease resistance proteins via helper NLR bioengineering Mauricio P. Contreras , Hsuan Pai , Muniyandi Selvaraj , Amirali Toghani , David M. Lawson , Yasin Tumtas , Cian Duggan , Enoch Lok Him Yuen , Clare E. M. Stevenson , Adeline Harant , Abbas Maqbool , Chih-Hang Wu , Tolga O. Bozkurt , Sophien Kamoun , Lida Derevnina Science Advances 9 DOI: 10.1126/sciadv.adg3861 Diamond Proposal Number(s): [18565] Open Access Show Abstract ▼ Abstract: Parasites counteract host immunity by suppressing helper nucleotide binding and leucine-rich repeat (NLR) proteins that function as central nodes in immune receptor networks. Understanding the mechanisms of immunosuppression can lead to strategies for bioengineering disease resistance. Here, we show that a cyst nematode virulence effector binds and inhibits oligomerization of the helper NLR protein NRC2 by physically preventing intramolecular rearrangements required for activation. An amino acid polymorphism at the binding interface between NRC2 and the inhibitor is sufficient for this helper NLR to evade immune suppression, thereby restoring the activity of multiple disease resistance genes. This points to a potential strategy for resurrecting disease resistance in crop genomes.	May 2023
I04-Macromolecular Crystallography	New dual inhibitors of bacterial topoisomerases with broad-spectrum antibacterial activity and in vivo efficacy against vancomycin-intermediate Staphylococcus aureus Martina Durcik , Andrej Emanuel Cotman , Žan Toplak , Štefan Možina , Žiga Skok , Petra Eva Szili , Márton Czikkely , Elvin Maharramov , Thu Hien Vu , Maria Vittoria Piras , Nace Zidar , Janez Ilaš , Anamarija Zega , Jurij Trontelj , Luis A. Pardo , Diarmaid Hughes , Douglas Huseby , Tália Berruga-Fernández , Sha Cao , Ivailo Simoff , Richard Svensson , Sergiy V. Korol , Zhe Jin , Francisca Vicente , Maria C. Ramos , Julia E. A. Mundy , Anthony Maxwell , Clare E. M. Stevenson , David M. Lawson , Björn Glinghammar , Eva Sjöström , Martin Bohlin , Joanna Oreskär , Sofie Alvér , Guido V. Janssen , Geert Jan Sterk , Danijel Kikelj , Csaba Pal , Tihomir Tomašič , Lucija Peterlin Mašič Journal Of Medicinal Chemistry 1703 DOI: 10.1021/acs.jmedchem.2c01905 Diamond Proposal Number(s): [25108] Open Access Show Abstract ▼ Abstract: A new series of dual low nanomolar benzothiazole inhibitors of bacterial DNA gyrase and topoisomerase IV were developed. The resulting compounds show excellent broad-spectrum antibacterial activities against Gram-positive Enterococcus faecalis, Enterococcus faecium and multidrug resistant (MDR) Staphylococcus aureus strains [best compound minimal inhibitory concentrations (MICs): range, <0.03125–0.25 μg/mL] and against the Gram-negatives Acinetobacter baumannii and Klebsiella pneumoniae (best compound MICs: range, 1–4 μg/mL). Lead compound 7a was identified with favorable solubility and plasma protein binding, good metabolic stability, selectivity for bacterial topoisomerases, and no toxicity issues. The crystal structure of 7a in complex with Pseudomonas aeruginosa GyrB24 revealed its binding mode at the ATP-binding site. Expanded profiling of 7a and 7h showed potent antibacterial activity against over 100 MDR and non-MDR strains of A. baumannii and several other Gram-positive and Gram-negative strains. Ultimately, in vivo efficacy of 7a in a mouse model of vancomycin-intermediate S. aureus thigh infection was also demonstrated.	Mar 2023
I04-Macromolecular Crystallography	The CTP-binding domain is disengaged from the DNA-binding domain in a co-crystal structure of Bacillus subtilis Noc-DNA complex Kirill V. Sukhoverkov , Adam S. B. Jalal , James R. Ault , Frank Sobott , David M. Lawson , Tung B. K. Le Journal Of Biological Chemistry 13 DOI: 10.1016/j.jbc.2023.103063 Diamond Proposal Number(s): [18565] Open Access Show Abstract ▼ Abstract: In Bacillus subtilis, a ParB-like nucleoid occlusion protein (Noc) binds specifically to Noc-binding sites (NBS) on the chromosome to help coordinate chromosome segregation and cell division. Noc does so by binding to cytidine triphosphate (CTP) to form large membrane-associated nucleoprotein complexes to physically inhibit the assembly of the cell division machinery. The site-specific binding of Noc to NBS DNA is a prerequisite for CTP-binding and the subsequent formation of a membrane-active DNA-entrapped protein complex. Here, we solve the structure of a C-terminally truncated B. subtilis Noc bound to NBS DNA to reveal the conformation of Noc at this crucial step. Our structure reveals the disengagement between the N-terminal CTP-binding domain and the NBS-binding domain of each DNA-bound Noc subunit, this is driven, in part, by the swapping of helices 4 and 5 at the interface of the two domains. Site-specific crosslinking data suggest that this conformation of Noc-NBS exists in solution. Overall, our results lend support to the recent proposal that parS/NBS-binding catalyzes CTP-binding and DNA-entrapment by preventing the re-engagement of the CTP-binding domain and the DNA-binding domain from the same ParB/Noc subunit.	Feb 2023
I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength) I04-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	Discovery and hit-to-lead optimization of benzothiazole scaffold-based DNA gyrase inhibitors with potent activity against Acinetobacter baumannii and Pseudomonas aeruginosa Andrej Emanuel Cotman , Martina Durcik , Davide Benedetto Tiz , Federica Fulgheri , Daniela Secci , Maša Sterle , Štefan Možina , Žiga Skok , Nace Zidar , Anamarija Zega , Janez Ilaš , Lucija Peterlin Mašič , Tihomir Tomašič , Diarmaid Hughes , Douglas L. Huseby , Sha Cao , Linnéa Garoff , Talía Berruga Fernández , Paraskevi Giachou , Lisa Crone , Ivailo Simoff , Richard Svensson , Bryndis Birnir , Sergiy V. Korol , Zhe Jin , Francisca Vicente , Maria C. Ramos , Mercedes De La Cruz , Björn Glinghammar , Lena Lenhammar , Sara R. Henderson , Julia E. A. Mundy , Anthony Maxwell , Claren E. M. Stevenson , David M. Lawson , Guido V. Janssen , Geert Jan Sterk , Danijel Kikelj Journal Of Medicinal Chemistry DOI: 10.1021/acs.jmedchem.2c01597 Diamond Proposal Number(s): [18565, 25108] Open Access Show Abstract ▼ Abstract: We have developed compounds with a promising activity against Acinetobacter baumannii and Pseudomonas aeruginosa, which are both on the WHO priority list of antibiotic-resistant bacteria. Starting from DNA gyrase inhibitor 1, we identified compound 27, featuring a 10-fold improved aqueous solubility, a 10-fold improved inhibition of topoisomerase IV from A. baumannii and P. aeruginosa, a 10-fold decreased inhibition of human topoisomerase IIα, and no cross-resistance to novobiocin. Cocrystal structures of 1 in complex with Escherichia coli GyrB24 and (S)-27 in complex with A. baumannii GyrB23 and P. aeruginosa GyrB24 revealed their binding to the ATP-binding pocket of the GyrB subunit. In further optimization steps, solubility, plasma free fraction, and other ADME properties of 27 were improved by fine-tuning of lipophilicity. In particular, analogs of 27 with retained anti-Gram-negative activity and improved plasma free fraction were identified. The series was found to be nongenotoxic, nonmutagenic, devoid of mitochondrial toxicity, and possessed no ion channel liabilities.	Jan 2023

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