scholarly journals The structure of the proteinaceous inhibitor PliI fromAeromonas hydrophilain complex with its target lysozyme

2015 ◽  
Vol 71 (2) ◽  
pp. 344-351 ◽  
Author(s):  
Seppe Leysen ◽  
Joris M. Van Herreweghe ◽  
Kazunari Yoneda ◽  
Makoto Ogata ◽  
Taichi Usui ◽  
...  
Keyword(s):  
Structural Data ◽  
Gram Negative Bacteria ◽  
Conserved Residues ◽  
New Class ◽  
Rational Development ◽  
Lysozyme Inhibitor ◽  
Meretrix Lusoria ◽  
Binding Groove ◽  
Proteinaceous Inhibitor ◽  
Microbiological Data

Recent microbiological data have revealed that Gram-negative bacteria are able to protect themselves against the lytic action of host lysozymes by secreting proteinaceous inhibitors. Four distinct classes of such inhibitors have been discovered that specifically act against c-type, g-type and i-type lysozymes. Here, the 1.24 Å resolution crystal structure of the periplasmic i-type lysozyme inhibitor fromAeromonas hydrophila(PliI-Ah) in complex with the i-type lysozyme fromMeretrix lusoriais reported. The structure is the first to explain the inhibitory mechanism of the PliI family at the atomic level. A distinct `ridge' formed by three exposed PliI loops inserts into the substrate-binding groove of the lysozyme, resulting in a complementary `key–lock' interface. The interface is principally stabilized by the interactions made by the PliI-Ah residues Ser104 and Tyr107 belonging to the conserved SGxY motif, as well as by the other conserved residues Ser46 and Asp76. The functional importance of these residues is confirmed by inhibition assays with the corresponding point mutants of PliI-Ah. The accumulated structural data on lysozyme–inhibitor complexes from several classes indicate that in all cases an extensive interface of either a single or a double `key–lock' type is formed, resulting in highly efficient inhibition. These data provide a basis for the rational development of a new class of antibacterial drugs.

scholarly journals The Fragment-Based Development of a Benzofuran Hit as a New Class of Escherichia coli DsbA Inhibitors

Molecules ◽  
2019 ◽  
Vol 24 (20) ◽  
pp. 3756 ◽  
Author(s):  
Luke F. Duncan ◽  
Geqing Wang ◽  
Olga V. Ilyichova ◽  
Martin J. Scanlon ◽  
Begoña Heras ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dissociation Constants ◽  
Gram Negative Bacteria ◽  
Nmr Studies ◽  
X Ray ◽  
New Class ◽  
Disulfide Oxidoreductase ◽  
Hydrophobic Binding ◽  
Binding Groove ◽  
Fragment Based Drug Discovery

A fragment-based drug discovery approach was taken to target the thiol-disulfide oxidoreductase enzyme DsbA from Escherichia coli (EcDsbA). This enzyme is critical for the correct folding of virulence factors in many pathogenic Gram-negative bacteria, and small molecule inhibitors can potentially be developed as anti-virulence compounds. Biophysical screening of a library of fragments identified several classes of fragments with affinity to EcDsbA. One hit with high mM affinity, 2-(6-bromobenzofuran-3-yl)acetic acid (6), was chemically elaborated at several positions around the scaffold. X-ray crystal structures of the elaborated analogues showed binding in the hydrophobic binding groove adjacent to the catalytic disulfide bond of EcDsbA. Binding affinity was calculated based on NMR studies and compounds 25 and 28 were identified as the highest affinity binders with dissociation constants (KD) of 326 ± 25 and 341 ± 57 µM respectively. This work suggests the potential to develop benzofuran fragments into a novel class of EcDsbA inhibitors.

scholarly journals The antimicrobial potential of cannabidiol

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Mark A. T. Blaskovich ◽  
Angela M. Kavanagh ◽  
Alysha G. Elliott ◽  
Bing Zhang ◽  
Soumya Ramu ◽  
...  
Keyword(s):  
Bacterial Infections ◽  
Modern Medicine ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
New Class ◽  
Clostridioides Difficile ◽  
Excellent Activity ◽  
Induce Resistance ◽  
First Time

AbstractAntimicrobial resistance threatens the viability of modern medicine, which is largely dependent on the successful prevention and treatment of bacterial infections. Unfortunately, there are few new therapeutics in the clinical pipeline, particularly for Gram-negative bacteria. We now present a detailed evaluation of the antimicrobial activity of cannabidiol, the main non-psychoactive component of cannabis. We confirm previous reports of Gram-positive activity and expand the breadth of pathogens tested, including highly resistant Staphylococcus aureus, Streptococcus pneumoniae, and Clostridioides difficile. Our results demonstrate that cannabidiol has excellent activity against biofilms, little propensity to induce resistance, and topical in vivo efficacy. Multiple mode-of-action studies point to membrane disruption as cannabidiol’s primary mechanism. More importantly, we now report for the first time that cannabidiol can selectively kill a subset of Gram-negative bacteria that includes the ‘urgent threat’ pathogen Neisseria gonorrhoeae. Structure-activity relationship studies demonstrate the potential to advance cannabidiol analogs as a much-needed new class of antibiotics.

scholarly journals Insights into AMS/PCAT transporters from biochemical and structural characterization of a double Glycine motif protease

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Silvia C Bobeica ◽  
Shi-Hui Dong ◽  
Liujie Huo ◽  
Nuria Mazo ◽  
Martin I McLaughlin ◽  
...  
Keyword(s):  
Structural Data ◽  
Leader Peptide ◽  
Gram Negative Bacteria ◽  
Protease Domain ◽  
Peptide Recognition ◽  
Gram Positive Bacteria ◽  
Dual Functional ◽  
Signaling Peptides

The secretion of peptides and proteins is essential for survival and ecological adaptation of bacteria. Dual-functional ATP-binding cassette transporters export antimicrobial or quorum signaling peptides in Gram-positive bacteria. Their substrates contain a leader sequence that is excised by an N-terminal peptidase C39 domain at a double Gly motif. We characterized the protease domain (LahT150) of a transporter from a lanthipeptide biosynthetic operon in Lachnospiraceae and demonstrate that this protease can remove the leader peptide from a diverse set of peptides. The 2.0 Å resolution crystal structure of the protease domain in complex with a covalently bound leader peptide demonstrates the basis for substrate recognition across the entire class of such transporters. The structural data also provide a model for understanding the role of leader peptide recognition in the translocation cycle, and the function of degenerate, non-functional C39-like domains (CLD) in substrate recruitment in toxin exporters in Gram-negative bacteria.

scholarly journals Using mutagenesis to explore conserved residues in the RNA-binding groove of influenza A virus nucleoprotein for antiviral drug development

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Chia-Lin Liu ◽  
Hui-Chen Hung ◽  
Shou-Chen Lo ◽  
Ching-Hui Chiang ◽  
I-Jung Chen ◽  
...  
Keyword(s):  
Drug Development ◽  
Influenza A Virus ◽  
Influenza A ◽  
Rna Binding ◽  
Antiviral Drug ◽  
Conserved Residues ◽  
Binding Groove

scholarly journals Functional Relationships between the AcrA Hairpin Tip Region and the TolC Aperture Tip Region for the Formation of the Bacterial Tripartite Efflux Pump AcrAB-TolC

2010 ◽  
Vol 192 (17) ◽  
pp. 4498-4503 ◽  
Author(s):  
Hong-Man Kim ◽  
Yongbin Xu ◽  
Minho Lee ◽  
Shunfu Piao ◽  
Se-Hoon Sim ◽  
...  
Keyword(s):  
Functional Data ◽  
Molecular Basis ◽  
Pathogenic Bacteria ◽  
Essential Role ◽  
Efflux Pump ◽  
Gram Negative Bacteria ◽  
Conserved Residues ◽  
Membrane Fusion Protein ◽  
Functional Relationships

ABSTRACT Tripartite efflux pumps found in Gram-negative bacteria are involved in antibiotic resistance and toxic-protein secretion. In this study, we show, using site-directed mutational analyses, that the conserved residues located in the tip region of the α-hairpin of the membrane fusion protein (MFP) AcrA play an essential role in the action of the tripartite efflux pump AcrAB-TolC. In addition, we provide in vivo functional data showing that both the length and the amino acid sequence of the α-hairpin of AcrA can be flexible for the formation of a functional AcrAB-TolC pump. Genetic-complementation experiments further indicated functional interrelationships between the AcrA hairpin tip region and the TolC aperture tip region. Our findings may offer a molecular basis for understanding the multidrug resistance of pathogenic bacteria.

scholarly journals Membrane Protein Complex ExbB4-ExbD1-TonB1from Escherichia coli Demonstrates Conformational Plasticity

2015 ◽  
Vol 197 (11) ◽  
pp. 1873-1885 ◽  
Author(s):  
Aleksandr Sverzhinsky ◽  
Jacqueline W. Chung ◽  
Justin C. Deme ◽  
Lucien Fabre ◽  
Kristian T. Levey ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Iron Acquisition ◽  
Proton Translocation ◽  
Three Dimensional ◽  
Structural Data ◽  
Structural Features ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Content Type

ABSTRACTIron acquisition at the outer membrane (OM) of Gram-negative bacteria is powered by the proton motive force (PMF) of the cytoplasmic membrane (CM), harnessed by the CM-embedded complex of ExbB, ExbD, and TonB. Its stoichiometry, ensemble structural features, and mechanism of action are unknown. By panning combinatorial phage libraries, periplasmic regions of dimerization between ExbD and TonB were predicted. Using overexpression of full-length His6-taggedexbB-exbDand S-taggedtonB, we purified detergent-solubilized complexes of ExbB-ExbD-TonB fromEscherichia coli. Protein-detergent complexes of ∼230 kDa with a hydrodynamic radius of ∼6.0 nm were similar to previously purified ExbB4-ExbD2complexes. Significantly, they differed in electronegativity by native agarose gel electrophoresis. The stoichiometry was determined to be ExbB4-ExbD1-TonB1. Single-particle electron microscopy agrees with this stoichiometry. Two-dimensional averaging supported the phage display predictions, showing two forms of ExbD-TonB periplasmic heterodimerization: extensive and distal. Three-dimensional (3D) particle classification showed three representative conformations of ExbB4-ExbD1-TonB1. Based on our structural data, we propose a model in which ExbD shuttles a proton across the CM via an ExbB interprotein rearrangement. Proton translocation would be coupled to ExbD-mediated collapse of extended TonB in complex with ligand-loaded receptors in the OM, followed by repositioning of TonB through extensive dimerization with ExbD. Here we present the first report for purification of the ExbB-ExbD-TonB complex, molar ratios within the complex (4:1:1), and structural biology that provides insights into 3D organization.IMPORTANCEReceptors in the OM of Gram-negative bacteria allow entry of iron-bound siderophores that are necessary for pathogenicity. Numerous iron-acquisition strategies rely upon a ubiquitous and unique protein for energization: TonB. Complexed with ExbB and ExbD, the Ton system links the PMF to OM transport. Blocking iron uptake by targeting a vital nanomachine holds promise in therapeutics. Despite much research, the stoichiometry, structural arrangement, and molecular mechanism of the CM-embedded ExbB-ExbD-TonB complex remain unreported. Here we demonstratein vitroevidence of ExbB4-ExbD1-TonB1complexes. Using 3D EM, we reconstructed the complex in three conformational states that show variable ExbD-TonB heterodimerization. Our structural observations form the basis of a model for TonB-mediated iron acquisition.

scholarly journals Curative Treatment of Severe Gram-Negative Bacterial Infections by a New Class of Antibiotics Targeting LpxC

mBio ◽  
2017 ◽  
Vol 8 (4) ◽  
Author(s):  
Nadine Lemaître ◽  
Xiaofei Liang ◽  
Javaria Najeeb ◽  
Chul-Jin Lee ◽  
Marie Titecat ◽  
...  
Keyword(s):  
Bacterial Infections ◽  
Biosynthetic Pathway ◽  
Lipid A ◽  
Multidrug Resistant ◽  
Gram Negative Bacteria ◽  
Drug Resistant ◽  
Bubonic Plague ◽  
Gram Negative ◽  
New Class

ABSTRACT The infectious diseases caused by multidrug-resistant bacteria pose serious threats to humankind. It has been suggested that an antibiotic targeting LpxC of the lipid A biosynthetic pathway in Gram-negative bacteria is a promising strategy for curing Gram-negative bacterial infections. However, experimental proof of this concept is lacking. Here, we describe our discovery and characterization of a biphenylacetylene-based inhibitor of LpxC, an essential enzyme in the biosynthesis of the lipid A component of the outer membrane of Gram-negative bacteria. The compound LPC-069 has no known adverse effects in mice and is effective in vitro against a broad panel of Gram-negative clinical isolates, including several multiresistant and extremely drug-resistant strains involved in nosocomial infections. Furthermore, LPC-069 is curative in a murine model of one of the most severe human diseases, bubonic plague, which is caused by the Gram-negative bacterium Yersinia pestis. Our results demonstrate the safety and efficacy of LpxC inhibitors as a new class of antibiotic against fatal infections caused by extremely virulent pathogens. The present findings also highlight the potential of LpxC inhibitors for clinical development as therapeutics for infections caused by multidrug-resistant bacteria. IMPORTANCE The rapid spread of antimicrobial resistance among Gram-negative bacilli highlights the urgent need for new antibiotics. Here, we describe a new class of antibiotics lacking cross-resistance with conventional antibiotics. The compounds inhibit LpxC, a key enzyme in the lipid A biosynthetic pathway in Gram-negative bacteria, and are active in vitro against a broad panel of clinical isolates of Gram-negative bacilli involved in nosocomial and community infections. The present study also constitutes the first demonstration of the curative treatment of bubonic plague by a novel, broad-spectrum antibiotic targeting LpxC. Hence, the data highlight the therapeutic potential of LpxC inhibitors against a wide variety of Gram-negative bacterial infections, including the most severe ones caused by Y. pestis and by multidrug-resistant and extensively drug-resistant carbapenemase-producing strains. IMPORTANCE The rapid spread of antimicrobial resistance among Gram-negative bacilli highlights the urgent need for new antibiotics. Here, we describe a new class of antibiotics lacking cross-resistance with conventional antibiotics. The compounds inhibit LpxC, a key enzyme in the lipid A biosynthetic pathway in Gram-negative bacteria, and are active in vitro against a broad panel of clinical isolates of Gram-negative bacilli involved in nosocomial and community infections. The present study also constitutes the first demonstration of the curative treatment of bubonic plague by a novel, broad-spectrum antibiotic targeting LpxC. Hence, the data highlight the therapeutic potential of LpxC inhibitors against a wide variety of Gram-negative bacterial infections, including the most severe ones caused by Y. pestis and by multidrug-resistant and extensively drug-resistant carbapenemase-producing strains.

scholarly journals Functional and structural characterization of Serratia marcescens ExbB: determinants of the interaction with HasB/TonB

2021 ◽  
Author(s):  
Valérie Biou ◽  
Ricardo Jorge D Adaixo ◽  
Mohamed Chami ◽  
Pierre-Damien Coureux ◽  
Benoist Laurent ◽  
...  
Keyword(s):  
Serratia Marcescens ◽  
Molecular Motor ◽  
Opportunistic Pathogen ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
E Coli ◽  
New Class ◽  
Heme Acquisition ◽  
Shed Light

ExbBD is part of a cytoplasmic membrane molecular motor driven by the proton-motive force. It belongs to the larger family of motors involved in nutriment import across the outer membrane of Gram-negative bacteria (ExbBD), flagellar rotation (MotAB) or late steps of cell division in Gram-negative bacteria (TolQR). ExbB and ExbD are integral membrane proteins with three (ExbB) or one (ExbD) transmembrane segment. Here we have solved by single-particle cryo-EM the structures of ExbB alone and of the ExbB-ExbD complex of the opportunistic pathogen Serratia marcescens. ExbBSm alone behaves as a stable pentamer, and the complex displays the ExbB5-ExbD2 stoichiometry. This is similar to what has been observed for ExbB-ExbD complexes from Escherichia coli and Pseudomonas savastanoi as well as MotAB complexes from various species. We identified residues located in the first TM of ExbBSm and ExbBEc that are likely involved in the interaction with TonB/HasB and that are essential for function. ExbBSm has a ca. 40 residues long periplasmic extension absent in E. coli. Such long ExbBs are found in some Gammaproteobacteria, and several genera of Alphaproteobacteria. We show that this extension interacts with HasB, a dedicated TonB paralog from the heme acquisition system (Has) from S. marcescens. We also show that it is involved in heme acquisition via the Has system from S. marcescens. ExbBSm represents thus a new class of ExbB protein and our results shed light on the specificity determinants between the ExbB-ExbD complex and their associated TonB partners.

scholarly journals SilverSil: A New Class of Antibacterial Materials of Broad Scope

2019 ◽  
Author(s):  
Keren Trabelsi ◽  
Rosaria Ciriminna ◽  
Yael Albo ◽  
Mario Pagliaro
Keyword(s):  
Low Cost ◽  
Consumer Products ◽  
Gram Negative Bacteria ◽  
Antibacterial Materials ◽  
New Class ◽  
Broad Scope ◽  
Organically Modified ◽  
Excellent Health ◽  
Organically Modified Silica ◽  
Hospital School

Consisting of organically modified silica (ORMOSIL) physically doped with Ag nanoparticles, the SilverSil new class of antibacterial materials of broad scope reported herein shows remarkably high and stable activity against representative Gram-positive and Gram-negative bacteria. The low cost, ease of application and excellent health and environmental profile of SilverSil hybrid glassy coatings open the route to their widespread utilization across domestic, hospital, school, industrial and commercial environments and in consumer products.<br>

scholarly journals A novel class of polymorphic toxins in Bacteroidetes

2020 ◽  
Vol 3 (4) ◽  
pp. e201900631
Author(s):  
Biswanath Jana ◽  
Dor Salomon ◽  
Eran Bosis
Keyword(s):  
Gut Microbiota ◽  
Unknown Function ◽  
Secretion System ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
New Class ◽  
Terminal Domain ◽  
Domain Of Unknown Function ◽  
Type Ix Secretion System ◽  
Encoding Gene

Bacteroidetes are Gram-negative bacteria that are abundant in the environment as well as in the gut microbiota of animals. Many bacteroidetes encode large proteins containing an N-terminal domain of unknown function, named TANFOR. In this work, we show that TANFOR-containing proteins carry polymorphic C-terminal toxin domains with predicted antibacterial and anti-eukaryotic activities. We also show that a C-terminal domain that is prevalent in TANFOR-containing proteins represents a novel family of antibacterial DNase toxins, which we named BaCT (Bacteroidetes C-terminal Toxin). Finally, we discover that TANFOR-encoding gene neighborhoods are enriched with genes that encode substrates of the type IX secretion system (T9SS), which is involved in exporting proteins from the periplasm across the outer membrane. Based on these findings, we conclude that TANFOR-containing proteins are a new class of polymorphic toxins, and we hypothesize that they are T9SS substrates.

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