scholarly journals Development of Anti-Virulence Therapeutics against Mono-ADP-Ribosyltransferase Toxins

Toxins ◽  
2020 ◽  
Vol 13 (1) ◽  
pp. 16
Author(s):  
Miguel R. Lugo ◽  
Allan R. Merrill
Keyword(s):  
Host Cell ◽  
Pathogenic Bacteria ◽  
Multi Drug Resistance ◽  
Post Translational Modification ◽  
Enzyme Active Site ◽  
Catalytic Function ◽  
Small Molecule Therapeutics ◽  
Adp Ribosyltransferase ◽  
Critical Process ◽  
Target Activity

Mono-ADP-ribosyltransferase toxins are often key virulence factors produced by pathogenic bacteria as tools to compromise the target host cell. These toxins are enzymes that use host cellular NAD+ as the substrate to modify a critical macromolecule target in the host cell machinery. This post-translational modification of the target macromolecule (usually protein or DNA) acts like a switch to turn the target activity on or off resulting in impairment of a critical process or pathway in the host. One approach to stymie bacterial pathogens is to curtail the toxic action of these factors by designing small molecules that bind tightly to the enzyme active site and prevent catalytic function. The inactivation of these toxins/enzymes is targeted for the site of action within the host cell and small molecule therapeutics can function as anti-virulence agents by disarming the pathogen. This represents an alternative strategy to antibiotic therapy with the potential as a paradigm shift that may circumvent multi-drug resistance in the offending microbe. In this review, work that has been accomplished during the past two decades on this approach to develop anti-virulence compounds against mono-ADP-ribosyltransferase toxins will be discussed.

scholarly journals A Structural Approach to Anti-Virulence: A Discovery Pipeline

2021 ◽  
Vol 9 (12) ◽  
pp. 2514
Author(s):  
Michael McCarthy ◽  
Monica Goncalves ◽  
Hannah Powell ◽  
Blake Morey ◽  
Madison Turner ◽  
...  
Keyword(s):  
Host Cell ◽  
Virulence Factors ◽  
Pathogenic Bacteria ◽  
Bacterial Virulence ◽  
Structural Approach ◽  
Multi Drug Resistance ◽  
Enzyme Active Site ◽  
Catalytic Function ◽  
Molecule Inhibitor ◽  
High Resolution Structure

The anti-virulence strategy is designed to prevent bacterial virulence factors produced by pathogenic bacteria from initiating and sustaining an infection. One family of bacterial virulence factors is the mono-ADP-ribosyltransferase toxins, which are produced by pathogens as tools to compromise the target host cell. These toxins are bacterial enzymes that exploit host cellular NAD+ as the donor substrate to modify an essential macromolecule acceptor target in the host cell. This biochemical reaction modifies the target macromolecule (often protein or DNA) and functions in a binary fashion to turn the target activity on or off by blocking or impairing a critical process or pathway in the host. A structural biology approach to the anti-virulence method to neutralize the cytotoxic effect of these factors requires the search and design of small molecules that bind tightly to the enzyme active site and prevent catalytic function essentially disarming the pathogen. This method requires a high-resolution structure to serve as the model for small molecule inhibitor development, which illuminates the path to drug development. This alternative strategy to antibiotic therapy represents a paradigm shift that may circumvent multi-drug resistance in the offending microbe through anti-virulence therapy. In this report, the rationale for the anti-virulence structural approach will be discussed along with recent efforts to apply this method to treat honey bee diseases using natural products.

scholarly journals Several New Putative Bacterial ADP-Ribosyltransferase Toxins Are Revealed from In Silico Data Mining, Including the Novel Toxin Vorin, Encoded by the Fire Blight Pathogen Erwinia amylovora

Toxins ◽  
2020 ◽  
Vol 12 (12) ◽  
pp. 792
Author(s):  
Olivier Tremblay ◽  
Zachary Thow ◽  
A. Rod Merrill
Keyword(s):  
Host Cell ◽  
In Silico ◽  
Pathogenic Bacteria ◽  
Erwinia Amylovora ◽  
Genomic Sequence ◽  
Whooping Cough ◽  
Sequence Data ◽  
Mass Spectrometry Analysis ◽  
Spectrometry Analysis ◽  
Adp Ribosyltransferase

Mono-ADP-ribosyltransferase (mART) toxins are secreted by several pathogenic bacteria that disrupt vital host cell processes in deadly diseases like cholera and whooping cough. In the last two decades, the discovery of mART toxins has helped uncover the mechanisms of disease employed by pathogens impacting agriculture, aquaculture, and human health. Due to the current abundance of mARTs in bacterial genomes, and an unprecedented availability of genomic sequence data, mART toxins are amenable to discovery using an in silico strategy involving a series of sequence pattern filters and structural predictions. In this work, a bioinformatics approach was used to discover six bacterial mART sequences, one of which was a functional mART toxin encoded by the plant pathogen, Erwinia amylovora, called Vorin. Using a yeast growth-deficiency assay, we show that wild-type Vorin inhibited yeast cell growth, while catalytic variants reversed the growth-defective phenotype. Quantitative mass spectrometry analysis revealed that Vorin may cause eukaryotic host cell death by suppressing the initiation of autophagic processes. The genomic neighbourhood of Vorin indicated that it is a Type-VI-secreted effector, and co-expression experiments showed that Vorin is neutralized by binding of a cognate immunity protein, VorinI. We demonstrate that Vorin may also act as an antibacterial effector, since bacterial expression of Vorin was not achieved in the absence of VorinI. Vorin is the newest member of the mART family; further characterization of the Vorin/VorinI complex may help refine inhibitor design for mART toxins from other deadly pathogens.

Prospective Application of Aptamer-based Assays and Therapeutics in Bloodstream Infections

2020 ◽  
Vol 20 (10) ◽  
pp. 831-840
Author(s):  
Weibin Li
Keyword(s):  
Pathogenic Bacteria ◽  
Genetic Diagnosis ◽  
Bloodstream Infections ◽  
High Specificity ◽  
Peptide Sequence ◽  
High Morbidity ◽  
Specificity And Sensitivity ◽  
Prospective Application ◽  
Small Molecule Therapeutics

Sepsis is still a severe health problem worldwide with high morbidity and mortality. Blood bacterial culture remains the gold standard for the detection of pathogenic bacteria in bloodstream infections, but it is time-consuming, and both the sophisticated equipment and well-trained personnel are required. Immunoassays and genetic diagnosis are expensive and limited to specificity and sensitivity. Aptamers are single-stranded deoxyribonucleic acid (ssDNA) and ribonucleic acid (RNA) oligonucleotide or peptide sequence generated in vitro based on the binding affinity of aptamer-target by a process known as Systematic Evolution of Ligands by Exponential Enrichment (SELEX). By taking several advantages over monoclonal antibodies and other conventional small-molecule therapeutics, such as high specificity and affinity, negligible batch-to-batch variation, flexible modification and production, thermal stability, low immunogenicity and lack of toxicity, aptamers are presently becoming promising novel diagnostic and therapeutic agents. This review describes the prospective application of aptamerbased laboratory diagnostic assays and therapeutics for pathogenic bacteria and toxins in bloodstream infections.

scholarly journals ADP-ribosylation of integrin α7 modulates the binding of integrin α7β1 to laminin

2004 ◽  
Vol 385 (1) ◽  
pp. 309-317 ◽  
Author(s):  
Zhefeng ZHAO ◽  
Joanna GRUSZCZYNSKA-BIEGALA ◽  
Anna ZOLKIEWSKA
Keyword(s):  
C2c12 Myotubes ◽  
Post Translational Modification ◽  
Integrin Β1 ◽  
Adp Ribosylation ◽  
In Vitro Binding ◽  
Vitro Binding ◽  
C2c12 Skeletal Muscle Cells ◽  
Adp Ribosyltransferase

The extracellular domain of integrin α7 is ADP-ribosylated by an arginine-specific ecto-ADP-ribosyltransferase after adding exogenous NAD+ to intact C2C12 skeletal muscle cells. The effect of ADP-ribosylation on the structure or function of integrin α7β1 has not been explored. In the present study, we show that ADP-ribosylation of integrin α7 takes place exclusively in differentiated myotubes and that this post-translational modification modulates the affinity of α7β1 dimer for its ligand, laminin. ADP-ribosylation in the 37-kDa ‘stalk’ region of α7 that takes place at micromolar NAD+ concentrations increases the binding of the α7β1 dimer to laminin. Increased in vitro binding of integrin α7β1 to laminin after ADP-ribosylation of the 37-kDa fragment of α7 requires the presence of Mn2+ and it is not observed in the presence of Mg2+. In contrast, ADP-ribosylation of the 63-kDa N-terminal region comprising the ligand-binding site of α7 that occurs at approx. 100 μM NAD+ inhibits the binding of integrin α7β1 to laminin. Furthermore, incubation of C2C12 myotubes with NAD+ increases the expression of an epitope on integrin β1 subunit recognized by monoclonal antibody 9EG7. We discuss our results based on the current models of integrin activation. We also hypothesize that ADP-ribosylation may represent a mechanism of regulation of integrin α7β1 function in myofibres in vivo when the continuity of the membrane is compromised and NAD+ is available as a substrate for ecto-ADP-ribosylation.

The computational intervention of macrolide antibiotics in the treatment of COVID-19

2021 ◽  
Vol 27 ◽  
Author(s):  
Firoz Anwar ◽  
Hisham N. Altayb ◽  
Fahad A. Al-Abbasi ◽  
Vikas Kumar ◽  
Mohammad Amjad Kamal
Keyword(s):  
Binding Energy ◽  
Progeny Virus ◽  
Macrolide Antibiotics ◽  
Reference Drug ◽  
Angiotensin Converting Enzyme 2 ◽  
Receptor Proteins ◽  
Enzyme Active Site ◽  
Catalytic Function ◽  
Main Protease ◽  
Spike Proteins

: The spike (S) glycoprotein of SARS coronavirus (SARS-CoV-2) and human Angiotensin-converting enzyme 2 (ACE2), are both considered the key factors for the initiation of virus infection. The present work is an effort for a computational target to block the spike proteins (S) and ACE2 receptor proteins with Macrolide antibiotics like Azithromycin, (AZM), Clarithromycin (CLAM) and Erythromycin (ERY) along with RNA-dependent RNA polymerase (RdRp). These compounds were able to block the residues (Arg553, Arg555, and Ala558) surrounding the deep grove catalytic site (Val557) of RdRp and thus plays an important role in tight blocking of enzyme active site. Reference drug Remdesivir was used to compare the docking score of antibiotics with RdRp. Docking value exhibited good binding energy (-7.7 up to -8.2 kcal/mol) with RdRp, indicating their potential as a potent RdRp inhibitor. Interaction of CLAM and ERY presented low binding energy (-6.8 and -6.6) with the ACE2 receptor. At the same time, CLAM exhibited a good binding affinity of -6.4 kcal/mol, making it an excellent tool to block the attachment of spike protein to ACE2 receptors. Macrolides not only affected the attachment to ACE2 but also blocked the spike proteins further, consequently inhibiting the internalization in the host cell. Three Alkyl bonds between Arg555, Ala558, and Met542 by CLAM and two Alkyl bonds of Arg624 and Lys621 by ERY plays an important role for RdRp inactivation that can prevent the rise of newly budded progeny virus. These macrolides interacted with the main protease protein in the pocket responsible for the dimerization and catalytic function of this protein. The interaction occurred with residue Glu166, along with the catalytic residues (Tyr343, and His235) of Endoribonuclease (NSP15) protein. The present study gives three-way options either by blocking S proteins or ACE2 receptor proteins or inhibiting RdRp to counter any effect of COVID-19 by macrolide and could be useful in the treatment of COVID-19 till some better option available.

scholarly journals Effects of O-GlcNAcylation on functional mitochondrial transfer from astrocytes

2020 ◽  
pp. 0271678X2096958 ◽  
Author(s):  
Ji-Hyun Park ◽  
Yoshihiko Nakamura ◽  
Wenlu Li ◽  
Gen Hamanaka ◽  
Ken Arai ◽  
...  
Keyword(s):  
Protein Modification ◽  
Mitochondrial Protein ◽  
Brefeldin A ◽  
Glucose Deprivation ◽  
Oxygen Glucose Deprivation ◽  
Loss Of Function ◽  
Post Translational Modification ◽  
Mitochondrial Transfer ◽  
The Central Nervous System ◽  
Critical Process

Mitochondria may be transferred from cell to cell in the central nervous system and this process may help defend neurons against injury and disease. But how mitochondria maintain their functionality during the process of release into extracellular space remains unknown. Here, we report that mitochondrial protein O-GlcNAcylation is a critical process to support extracellular mitochondrial functionality. Activation of CD38-cADPR signaling in astrocytes robustly induced protein O-GlcNAcylation in mitochondria, while oxygen-glucose deprivation and reoxygenation showed transient and mild protein modification. Blocking the endoplasmic reticulum – Golgi trafficking with Brefeldin A or slc35B4 siRNA reduced O-GlcNAcylation, and resulted in the secretion of mitochondria with decreased membrane potential and mtDNA. Finally, loss-of-function studies verified that O-GlcNAc-modified mitochondria demonstrated higher levels of neuroprotection after astrocyte-to-neuron mitochondrial transfer. Collectively, these findings suggest that post-translational modification by O-GlcNAc may be required for supporting the functionality and neuroprotective properties of mitochondria released from astrocytes.

A DFT study on the catalytic mechanism of UDP-glucose dehydrogenase

2010 ◽  
Vol 88 (8) ◽  
pp. 804-814 ◽  
Author(s):  
WenJuan Huang ◽  
Jorge Llano ◽  
James W. Gauld
Keyword(s):  
Pathogenic Bacteria ◽  
Catalytic Mechanism ◽  
Glucuronic Acid ◽  
Glucose Dehydrogenase ◽  
Polarizable Continuum Model ◽  
Uridine Diphosphate ◽  
Reaction Field ◽  
Enzyme Active Site ◽  
Site Model ◽  
Diphosphate Glucose

Uridine 5′-diphosphate glucuronic acid (UDPGlcUA) is a key intermediary metabolite in many species, including pathogenic bacteria and humans. It is biosynthesized from UDP-glucose (UDPGlc) by uridine diphosphate glucose dehydrogenase (UDPGlcDH) via a twofold two-electron–one-proton oxidation that successively transforms the 6-hydroxymethyl of glucopyranose into a formyl, and the latter into the final carboxylic function. The catalytic mechanism of UDPGlcDH was investigated using a large enzyme active-site model in combination with the B3LYP method and the polarizable continuum model (IEF-PCM) self-consistent reaction field. The latter was used to correct for the long-range electrostatic effect of the protein environment. The overall mechanism consists of four catalytic steps: (i) NAD+-dependent oxidation of glucose to glucuronaldehyde, (ii) nucleophilic addition of Cys260–SH to glucuronaldehyde to form a 6-thiohemiacetal intermediate, (iii) NAD+-dependent oxidation of the 6-thiohemiacetal to form a 6-thioester intermediate, and finally, (iv) hydrolysis of the 6-thioester to give glucuronic acid. In addition, this study also provides insight into the debated roles of Lys204 and Asp264, and the most likely protonation state of a reactive Michaelis complex of UDPGlcDH.

scholarly journals The Sinorhizobium (Ensifer) fredii HH103 Nodulation Outer Protein NopI Is a Determinant for Efficient Nodulation of Soybean and Cowpea Plants

2016 ◽  
Vol 83 (5) ◽  
Author(s):  
Irene Jiménez-Guerrero ◽  
Francisco Pérez-Montaño ◽  
Carlos Medina ◽  
Francisco Javier Ollero ◽  
Francisco Javier López-Baena
Keyword(s):  
Adenylate Cyclase ◽  
Host Cell ◽  
Host Range ◽  
Pathogenic Bacteria ◽  
Defense Responses ◽  
Host Cells ◽  
Broad Host Range ◽  
Sinorhizobium Fredii ◽  
Symbiotic Efficiency ◽  
Content Type

ABSTRACT The type III secretion system (T3SS) is a specialized secretion apparatus that is commonly used by many plant and animal pathogenic bacteria to deliver proteins, termed effectors, to the interior of the host cells. These effectors suppress host defenses and interfere with signal transduction pathways to promote infection. Some rhizobial strains possess a functional T3SS, which is involved in the suppression of host defense responses, host range determination, and symbiotic efficiency. The analysis of the genome of the broad-host-range rhizobial strain Sinorhizobium fredii HH103 identified eight genes that code for putative T3SS effectors. Three of these effectors, NopL, NopP, and NopI, are Rhizobium specific. In this work, we demonstrate that NopI, whose amino acid sequence shows a certain similarity with NopP, is secreted through the S. fredii HH103 T3SS in response to flavonoids. We also determined that NopL can be considered an effector since it is directly secreted to the interior of the host cell as demonstrated by adenylate cyclase assays. Finally, the symbiotic phenotype of single, double, and triple nopI, nopL, and nopP mutants in soybean and cowpea was assayed, showing that NopI plays an important role in determining the number of nodules formed in both legumes and that the absence of both NopL and NopP is highly detrimental for symbiosis. IMPORTANCE The paper is focused on three Rhizobium-specific T3SS effectors of Sinorhizobium fredii HH103, NopL, NopP, and NopI. We demonstrate that S. fredii HH103 is able to secrete through the T3SS in response to flavonoids the nodulation outer protein NopI. Additionally, we determined that NopL can be considered an effector since it is secreted to the interior of the host cell as demonstrated by adenylate cyclase assays. Finally, nodulation assays of soybean and cowpea indicated that NopI is important for the determination of the number of nodules formed and that the absence of both NopL and NopP negatively affected nodulation.

scholarly journals Multi–drug resistant Staphylococcus aureus isolated from milk, chicken meat, beef and egg in Bangladesh

2018 ◽  
Vol 5 (2) ◽  
pp. 175-183 ◽  
Author(s):  
Mohammad Anisur Rahman ◽  
AKM Anisur Rahman ◽  
Md Ariful Islam ◽  
Md Mahbub Alam
Keyword(s):  
Pathogenic Bacteria ◽  
Dairy Farm ◽  
Food Samples ◽  
Staphylococcal Infection ◽  
Chicken Meat ◽  
Multi Drug Resistance ◽  
Highly Sensitive ◽  
Antimicrobial Susceptibility Tests ◽  
Susceptibility Tests ◽  
Rational Use Of Antibiotics

Staphylococcal infection is one of the most common food-borne diseases in the world. Moreover, antimicrobial resistance of pathogenic bacteria, including Staphylococcus (S.) aureus is an emerging problem of food safety. This study was conducted to investigate the prevalence of S. aureus in milk, chicken meat, egg and beef; and to determine the multi-drug resistance (MDR) profile of S. aureus in Mymensingh and Gazipur districts, Bangladesh. A total of 189 samples of milk (n=108), chicken meat (n=51), egg (n=20) and beef (n=10) were collected from Bangladesh Agricultural University dairy farm, American dairy farm, Gazipur and different dairy farms of municipal area and retail shops during July 2016 to June 2018. S. aureus were isolated and identified by conventional methods and polymerase chain reaction (PCR). Antimicrobial susceptibility tests were done through disc diffusion test using 10 commonly used antibiotics. The overall prevalence of S. aureus in all food samples was 43.39%. A total of 39 (76.47%) chicken meat, 25 (23.15%) milk, 11(55%) egg and 07 (70%) beef samples were S. aureus positive through conventional method. Among 82 culture positive samples only 39 samples (47.56%) were confirmed by PCR. Antibiogram study showed that S. aureus isolated from chicken meat were mostly resistant to oxytetracycline (71.79%); and highly sensitive to amikacin (100%) and neomycin (100%). S. aureus isolated from milk samples were highly sensitive to neomycin (100%), and resistant to amikacin (56%). Only 28.57% isolates of S. aureus originated from beef samples were resistant to oxytetracycline and 100 % isolates were sensitive to ciprofloxacin, gentamicin, erythromycin, azithromycin, doxycycline. Similarly, S. aureus isolated from egg samples were resistant to erythromycin (81.82%) and 100% sensitive to amikacin. Out of 41.46% MDR isolates 12%, 53.85%, 90.91% and 0% of the S. aureus originated from milk, chicken meat, egg and beef respectively. The higher prevalence of S. aureus in chicken meat, beef, egg and milk indicates unhygienic production, marketing and processing of these foods. Presence of MDR S. aureus in these foods might pose serious public health threats. Rational use of antibiotics with higher sensitivity should be prescribed in managing poultry diseases to reduce re-emerging MDR in Bangladesh.Res. Agric., Livest. Fish.5(2): 175-183, August 2018

scholarly journals Recruitment of Mammalian Cell Fibronectin to the Surface of Chlamydia trachomatis

2002 ◽  
Vol 70 (7) ◽  
pp. 3935-3938 ◽  
Author(s):  
Betsy J. Kleba ◽  
Erin Banta ◽  
Erika A. Lindquist ◽  
Richard S. Stephens
Keyword(s):  
Chlamydia Trachomatis ◽  
Heparan Sulfate ◽  
Host Cell ◽  
Mammalian Cell ◽  
Pathogenic Bacteria ◽  
Cell Binding ◽  
Cell Functions ◽  
Wide Range

ABSTRACT Pathogenic bacteria exploit the presence of various host cell molecules in order to colonize new tissues. Fibronectin is involved in a wide range of cell functions in vivo, and staphylococci, streptococci, and gonococci have evolved mechanisms to utilize this glycoprotein to mediate host cell binding. We show that elementary bodies (EB) from two biovars of Chlamydia trachomatis recruit fibronectin to their surfaces upon lysis of the host cell. We also demonstrate that a heparan sulfate lyase-sensitive molecule on chlamydial EB is responsible for binding at least a portion of this fibronectin.

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