scholarly journals Brevinin-2GHk from Sylvirana guentheri and the Design of Truncated Analogs Exhibiting the Enhancement of Antimicrobial Activity

Antibiotics ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 85 ◽  
Author(s):  
Guanzhu Chen ◽  
Yuxi Miao ◽  
Chengbang Ma ◽  
Mei Zhou ◽  
Zhanzhong Shi ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Mammalian Cells ◽  
Structural Changes ◽  
Structural Characteristics ◽  
Helical Conformation ◽  
Skin Secretion ◽  
Mature Peptide ◽  
Diverse Range ◽  
Low Cytotoxicity ◽  
Α Helix

Brevinins are an important antimicrobial peptide (AMP) family discovered in the skin secretions of Ranidae frogs. The members demonstrate a typical C-terminal ranabox, as well as a diverse range of other structural characteristics. In this study, we identified a novel brevinin-2 peptide from the skin secretion of Sylvirana guentheri, via cloning transcripts, and identifying the expressed mature peptide, in the skin secretion. The confirmed amino acid sequence of the mature peptide was designated brevinin-2GHk (BR2GK). Moreover, as a previous study had demonstrated that the N-terminus of brevinin-2 is responsible for exerting antimicrobial activity, we also designed a series of truncated derivatives of BR2GK. The results show that the truncated derivatives exhibit significantly improved antimicrobial activity and cytotoxicity compared to the parent peptide, except a Pro14 substituted analog. The circular dichroism (CD) analysis of this analog revealed that it did not fold into a helical conformation in the presence of either lipopolysaccharides (LPS) or TFE, indicating that position 14 is involved in the formation of the α-helix. Furthermore, three more analogs with the substitutions of Ala, Lys and Arg at the position 14, respectively, revealed the influence on the membrane disruption potency on bacteria and mammalian cells by the structural changes at this position. Overall, the N-terminal 25-mer truncates demonstrated the potent antimicrobial activity with low cytotoxicity.

scholarly journals Antimicrobial Peptide CMA3 Derived from the CA-MA Hybrid Peptide: Antibacterial and Anti-inflammatory Activities with Low Cytotoxicity and Mechanism of Action in Escherichia coli

2015 ◽  
Vol 60 (1) ◽  
pp. 495-506 ◽  
Author(s):  
Jong-kook Lee ◽  
Chang Ho Seo ◽  
Tudor Luchian ◽  
Yoonkyung Park
Keyword(s):  
Escherichia Coli ◽  
Antimicrobial Activity ◽  
Antimicrobial Peptide ◽  
Blood Cells ◽  
Mammalian Cells ◽  
White Blood Cells ◽  
Raw 264.7 Cells ◽  
Content Type ◽  
Anti Inflammatory ◽  
Low Cytotoxicity

ABSTRACTCA-MA is a hybrid antimicrobial peptide (AMP) derived from two naturally occurring AMPs, cecropin A and magainin 2. CA-MA shows strong antimicrobial activity against Gram-negative and Gram-positive bacteria but also exhibits cytotoxicity toward mammalian cells. Our objective was to identify CA-MA analogues with reduced cytotoxicity by systematic replacement of amino acids with positively charged R groups (His and Lys), aliphatic R groups (Leu), or polar R groups (Glu). Among the CA-MA analogues studied (CMA1 to -6), CMA3 showed the strongest antimicrobial activity, including against drug-resistantEscherichia coliandPseudomonas aeruginosastrains isolated from hospital patients. CMA3 appeared to act by inducing pore formation (toroidal model) in the bacterial membrane. In cytotoxicity assays, CMA3 showed little cytotoxicity toward human red blood cells (hRBCs) or HaCaT cells. Additionally, no fluorescence was released from small or giant unilamellar vesicles exposed to 60 μM CMA3 for 80 s, whereas fluorescence was released within 35 s upon exposure to CA-MA. CMA3 also exerted strong lipopolysaccharide (LPS)-neutralizing activity in RAW 264.7 cells, and BALB/c mice exposed to LPS after infection byEscherichia colishowed improved survival after administration of one 0.5-mg/kg of body weight or 1-mg/kg dose of CMA3. Finally, in a mouse model of septic shock, CMA3 reduced the levels of proinflammatory factors, including both nitric oxide and white blood cells, and correspondingly reduced lung tissue damage. This study suggests that CMA3 is an antimicrobial/antiendotoxin peptide that could serve as the basis for the development of anti-inflammatory and/or antimicrobial agents with low cytotoxicity.

scholarly journals Stapled Anoplin as an Antibacterial Agent

2021 ◽  
Vol 12 ◽  
Author(s):  
Monika Wojciechowska ◽  
Julia Macyszyn ◽  
Joanna Miszkiewicz ◽  
Renata Grzela ◽  
Joanna Trylska
Keyword(s):  
Antimicrobial Activity ◽  
Helical Structure ◽  
Eukaryotic Cells ◽  
Helical Conformation ◽  
Iodide Uptake ◽  
Gram Positive ◽  
Amphipathic Peptide ◽  
Gram Negative ◽  
Gram Positive Bacteria ◽  
Α Helix

Anoplin is a linear 10-amino acid amphipathic peptide (Gly-Leu-Leu-Lys-Arg-Ile-Lys-Thr-Leu-Leu-NH2) derived from the venom sac of the solitary wasp. It has broad antimicrobial activity, including an antibacterial one. However, the inhibition of bacterial growth requires several dozen micromolar concentrations of this peptide. Anoplin is positively charged and directly interacts with anionic biological membranes forming an α-helix that disrupts the lipid bilayer. To improve the bactericidal properties of anoplin by stabilizing its helical structure, we designed and synthesized its analogs with hydrocarbon staples. The staple was introduced at two locations resulting in different charges and amphipathicity of the analogs. Circular dichroism studies showed that all modified anoplins adopted an α-helical conformation, both in the buffer and in the presence of membrane mimics. As the helicity of the stapled anoplins increased, their stability in trypsin solution improved. Using the propidium iodide uptake assay in Escherichia coli and Staphylococcus aureus, we confirmed the bacterial membrane disruption by the stapled anoplins. Next, we tested the antimicrobial activity of peptides on a range of Gram-negative and Gram-positive bacteria. Finally, we evaluated peptide hemolytic activity on sheep erythrocytes and cytotoxicity on human embryonic kidney 293 cells. All analogs showed higher antimicrobial activity than unmodified anoplin. Depending on the position of the staple, the peptides were more effective either against Gram-negative or Gram-positive bacteria. Anoplin[5-9], with a lower positive charge and increased hydrophobicity, had higher activity against Gram-positive bacteria but also showed hemolytic and destructive effects on eukaryotic cells. Contrary, anoplin[2-6] with a similar charge and amphipathicity as natural anoplin effectively killed Gram-negative bacteria, also pathogenic drug-resistant strains, without being hemolytic and toxic to eukaryotic cells. Our results showed that anoplin charge, amphipathicity, and location of hydrophobic residues affect the peptide destructive activity on the cell wall, and thus, its antibacterial activity. This means that by manipulating the charge and position of the staple in the sequence, one can manipulate the antimicrobial activity.

Effect of ultrasound on the structural characteristics of fresh skim milk

2019 ◽  
Vol 26 (3) ◽  
pp. 222-230
Author(s):  
Jitao Yang ◽  
Min Yang ◽  
Juanjuan Qin ◽  
Qiubing Zeng ◽  
Yucheng Wang ◽  
...  
Keyword(s):  
Particle Size ◽  
Zeta Potential ◽  
Structural Changes ◽  
Structural Characteristics ◽  
Skim Milk ◽  
Fluorescence Properties ◽  
Net Charge ◽  
Acid Sodium Salt ◽  
Α Helix ◽  
Β Sheet

The structural changes of skim milk caused by sonication were evaluated by particle size, zeta-potential, turbidity, scanning electron microscopy, Fourier transform infrared spectroscopy, and intrinsic and 8-anilino-1-naphthalenesulfonic acid sodium salt fluorescence properties. The results showed that the particle size and zeta-potential of skim milk remained constant with 1 min ultrasonication, and increased significantly when the duration of sonication was extended to 3 min. With 3–10 min ultrasonic treatment, the diameter and net charge of particles in skim milk changed scarcely. According to the topography, the integrity of casein micelles was not damaged by 30 min sonication, but the turbidity decreased sharply with sonication above 5 min. The secondary structure of protein in skim milk changed after 1 min sonication, shown by a significant increase of α-helix content and decrease in the irregularity of β-sheet. The intrinsic fluorescence intensity of skim milk with 1 min sonication increased dramatically with a shift in the maximum emission wavelength. The fluorescence properties revealed that the spatial structure of protein in skim milk changed by sonication.

scholarly journals Wide-Antimicrobial Spectrum of Picolinium Salts

Molecules ◽  
2020 ◽  
Vol 25 (9) ◽  
pp. 2254 ◽  
Author(s):  
Sarka Salajkova ◽  
Marketa Benkova ◽  
Jan Marek ◽  
Radek Sleha ◽  
Lukas Prchal ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Biological Activity ◽  
Mammalian Cells ◽  
Alkyl Chain ◽  
Critical Micellar Concentration ◽  
Antimicrobial Compounds ◽  
Methyl Substituent ◽  
Antimicrobial Spectrum ◽  
Alkyl Chains ◽  
Low Cytotoxicity

Nosocomial infections, which greatly increase morbidity among hospitalized patients, together with growing antibiotic resistance still encourage many researchers to search for novel antimicrobial compounds. Picolinium salts with different lengths of alkyl chains (C12, C14, C16) were prepared by Menshutkin-like reaction and evaluated with respect to their biological activity, i.e., lipophilicity and critical micellar concentration. Picolinium salts with C14 and C16 side chains achieved similar or even better results when in terms of antimicrobial efficacy than benzalkoniums; notably, their fungicidal efficiency was substantially more potent. The position of the methyl substituent on the aromatic ring does not seem to affect antimicrobial activity, in contrast to the effect of length of the N-alkyl chain. Concurrently, picolinium salts exhibited satisfactory low cytotoxicity against mammalian cells, i.e., lower than that of benzalkonium compounds, which are considered as safe.

A Comparative Study of the Antimicrobial and Structural Properties of Short Peptides and Lipopeptides Containing a Repetitive Motif KLFK

2019 ◽  
Vol 26 (3) ◽  
pp. 192-203
Author(s):  
María Verónica Húmpola ◽  
María Carolina Rey ◽  
Pablo Gabriel Spontón ◽  
Arturo Carlos Simonetta ◽  
Georgina Guadalupe Tonarelli
Keyword(s):  
Antimicrobial Activity ◽  
Secondary Structure ◽  
Mammalian Cells ◽  
Therapeutic Index ◽  
Dilution Method ◽  
Hemolysis Assay ◽  
Improved Stability ◽  
Repetitive Motif ◽  
Bacteria And Fungi ◽  
Α Helix

Background:In the last years, Antimicrobial Peptides (AMPs) and lipopeptides have received attention as promising candidates to treat infections caused by resistant microorganisms. </P><P> Objective: The main objective of this study was to investigate the effect of repetitive KLFK motifs and the attachment of aliphatic acids to the N-terminus of (KLFK)n peptides on therapeutic properties.Methods:Minimal inhibitory concentration against Gram (+) and (-) bacteria and yeast of synthetic compounds were determined by broth microtiter dilution method, and the toxicity was evaluated by hemolysis assay. Membrane-peptide interaction studies were performed with model phospholipid membranes mimicking those of bacterial and mammalian cells by Fluorescence Spectroscopy. The secondary structure in solution and membranes was determined by Circular Dichroism.Results:Our results showed that the resulting compounds have inhibitory activity against bacteria and fungi. The (KLFK)3 peptide showed the highest therapeutic index against bacterial and yeast strains, and the (KLFK)2 peptide conjugated with octanoic acid was the most active against yeasts. All the lipopeptides containing long-chain fatty acids (C14 or longer) were highly hemolytic at low concentrations. The antimicrobial activity of (KLFK)2 and (KLFK)3 lipopeptides was mainly associated with improved stability of the amphipathic secondary structure, which showed high contributions of α-helix in dipalmitoylphosphatidylglycerol (DPPG) vesicles.Conclusion:The repetition of the KLFK sequence and the conjugation with lipid tails allowed obtained compounds with high antimicrobial activity and low toxicity, becoming good candidates for treating infectious diseases.

Synthesis and Structure−Activity Relationship Studies of N-monosubstituted Aroylthioureas as Urease Inhibitors

2020 ◽  
Vol 16 ◽  
Author(s):  
Wei-Wei Ni ◽  
Hai-Lian Fang ◽  
Ya-Xi Ye ◽  
Wei-Yi Li ◽  
Li Liu ◽  
...  
Keyword(s):  
Regression Analysis ◽  
Mammalian Cells ◽  
Intact Cell ◽  
Linear Regression Analysis ◽  
Positive Control ◽  
Acetohydroxamic Acid ◽  
Urease Inhibitors ◽  
Ic50 Value ◽  
Ic50 Values ◽  
Low Cytotoxicity

Background: Thiourea is a classical urease inhibitor usually as a positive control, and many N,N`-disubstituted thioureas have been determined as urease inhibitors. However, due to steric hindrance, N,N`-disubstituted thiourea motif could not bind urease as thiourea. On the contrary, N-monosubstituted thioureas with a tiny thiourea motif could theoretically bind into the active pocket as thiourea. Objective: A series of N-monosubstituted aroylthioureas were designed and synthesized for evaluation as urease inhibitors. Methods: Urease inhibition was determined by the indophenol method and IC50 values were calculated using computerized linear regression analysis of quantal log dose-probit functions. The kinetic parameters were estimated viasurface plasmon resonance (SPR) and by nonlinear regression analysis based on the mixed type inhibition model derived from Michaelis-Menten kinetics. Results: Compounds b2, b11and b19 reversibly inhibited urease with a mixed mechanism, and showed excellent potency against both cell-free urease and urease in intact cell, with IC50 values being 90-to 450-fold and 5-to 50-fold lower than the positive control acetohydroxamic acid, respectively. The most potent compound b11 showed IC50 value of 0.060 ±0.004μM against cell-free urease, which bound to urea binding site with a very low KDvalue (0.420±0.003nM) and a very long residence time (6.7 min). Compound b11was also demonstrated having very low cytotoxicity to mammalian cells. Conclusion: These results revealed that N-monosubstituted aroylthioureas clearly bind the active site of urease as expected, and represent a new class of urease inhibitors for the development of potential therapeutics against infections caused by ure-ase-containing pathogens.

scholarly journals Exploiting DNA Endonucleases to Advance Mechanisms of DNA Repair

Biology ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 530
Author(s):  
Marlo K. Thompson ◽  
Robert W. Sobol ◽  
Aishwarya Prakash
Keyword(s):  
Dna Repair ◽  
Mammalian Cells ◽  
Genome Stability ◽  
Gene Editing ◽  
Adaptive Immune System ◽  
Zinc Finger Nucleases ◽  
Specific Gene ◽  
Target Sequence ◽  
Transcription Activator ◽  
Low Cytotoxicity

The earliest methods of genome editing, such as zinc-finger nucleases (ZFN) and transcription activator-like effector nucleases (TALENs), utilize customizable DNA-binding motifs to target the genome at specific loci. While these approaches provided sequence-specific gene-editing capacity, the laborious process of designing and synthesizing recombinant nucleases to recognize a specific target sequence, combined with limited target choices and poor editing efficiency, ultimately minimized the broad utility of these systems. The discovery of clustered regularly interspaced short palindromic repeat sequences (CRISPR) in Escherichia coli dates to 1987, yet it was another 20 years before CRISPR and the CRISPR-associated (Cas) proteins were identified as part of the microbial adaptive immune system, by targeting phage DNA, to fight bacteriophage reinfection. By 2013, CRISPR/Cas9 systems had been engineered to allow gene editing in mammalian cells. The ease of design, low cytotoxicity, and increased efficiency have made CRISPR/Cas9 and its related systems the designer nucleases of choice for many. In this review, we discuss the various CRISPR systems and their broad utility in genome manipulation. We will explore how CRISPR-controlled modifications have advanced our understanding of the mechanisms of genome stability, using the modulation of DNA repair genes as examples.

scholarly journals Structural and Technological Approach to Reveal the Role of the Lipid Phase in the Formation of Soy Emulsion Gels with Chia Oil

Gels ◽  
2021 ◽  
Vol 7 (2) ◽  
pp. 48
Author(s):  
Ana M. Herrero ◽  
Claudia Ruiz-Capillas
Keyword(s):  
Raman Spectroscopy ◽  
Structural Properties ◽  
Structural Changes ◽  
Protein Secondary Structure ◽  
Lipid Phase ◽  
Α Helix ◽  
Β Sheet ◽  
Shed Light ◽  
Chia Oil

Considerable attention has been paid to emulsion gels (EGs) in recent years due to their interesting applications in food. The aim of this work is to shed light on the role played by chia oil in the technological and structural properties of EGs made from soy protein isolates (SPI) and alginate. Two systems were studied: oil-free SPI gels (SPI/G) and the corresponding SPI EGs (SPI/EG) that contain chia oil. The proximate composition, technological properties (syneresis, pH, color and texture) and structural properties using Raman spectroscopy were determined for SPI/G and SPI/EG. No noticeable (p > 0.05) syneresis was observed in either sample. The pH values were similar (p > 0.05) for SPI/G and SPI/EG, but their texture and color differed significantly depending on the presence of chia oil. SPI/EG featured significantly lower redness and more lightness and yellowness and exhibited greater puncture and gel strengths than SPI/G. Raman spectroscopy revealed significant changes in the protein secondary structure, i.e., higher (p < 0.05) α-helix and lower (p < 0.05) β-sheet, turn and unordered structures, after the incorporation of chia oil to form the corresponding SPI/EG. Apparently, there is a correlation between these structural changes and the textural modifications observed.

scholarly journals A “Target Class” Screen to Identify Activators of Two-Pore Domain Potassium (K2P) Channels

2020 ◽  
pp. 247255522097612
Author(s):  
David McCoull ◽  
Emma Ococks ◽  
Jonathan M. Large ◽  
David C. Tickle ◽  
Alistair Mathie ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Point Of View ◽  
Resting Membrane Potential ◽  
Small Molecule Activation ◽  
Target Class ◽  
Index Set ◽  
Diverse Range ◽  
K2p Channels ◽  
Additional Approach ◽  
Pore Domain

Two-pore domain potassium (K2P) channels carry background (or leak) potassium current and play a key role in regulating resting membrane potential and cellular excitability. Accumulating evidence points to a role for K2Ps in human pathophysiologies, most notably in pain and migraine, making them attractive targets for therapeutic intervention. However, there remains a lack of selective pharmacological tools. The aim of this work was to apply a “target class” approach to investigate the K2P superfamily and identify novel activators across all the described subclasses of K2P channels. Target class drug discovery allows for the leveraging of accumulated knowledge and maximizing synergies across a family of targets and serves as an additional approach to standard target-based screening. A common assay platform using baculovirus (BacMam) to transiently express K2P channels in mammalian cells and a thallium flux assay to determine channel activity was developed, allowing the simultaneous screening of multiple targets. Importantly, this system, by allowing precise titration of channel function, allows optimization to facilitate the identification of activators. A representative set of channels (THIK-1, TWIK-1, TREK-2, TASK-3, and TASK-2) were screened against a library of Food and Drug Administration (FDA)-approved compounds and the LifeArc Index Set. Activators were then analyzed in concentration–response format across all channels to assess selectivity. Using the target class approach to investigate the K2P channels has enabled us to determine which of the K2Ps are amenable to small-molecule activation, de-risk multiple channels from a technical point of view, and identify a diverse range of previously undescribed pharmacology.

Study of Properties of Expanded Graphite – Polymer Porous Composite by Acoustic Emission Method

2013 ◽  
Vol 58 (4) ◽  
pp. 1331-1336 ◽  
Author(s):  
J. Berdowski ◽  
S. Berdowska ◽  
F. Aubry
Keyword(s):  
Acoustic Emission ◽  
Structural Changes ◽  
Structural Characteristics ◽  
Physical And Mechanical Properties ◽  
Expanded Graphite ◽  
Composite Membranes ◽  
Acoustic Emission Method ◽  
Emission Method ◽  
Graphite Composite ◽  
Wide Range

Abstract The purpose of this paper was to investigate the physical and mechanical properties of compressed expanded graphite (CEG) and their porous derivatives after impregnation, polymerization; and carbonization by the use of acoustic emission method (AE). The mechanical and structural characteristics of compressed expanded graphite and their three groups of porous composites after each technological process are presented and discussed. The measurements of acoustic emission parameters in these materials were carried out at wide range of frequency of the waves (0.1÷2.5 MHz). The changes of two of parameters: - AE pulses counts rate and spectrum distribution of AE waves - are presented in this paper. The analysis of the respective parameters AE also gives possibility to determine the micro- and macro structural changes of materials at different levels of technological processes. Applications of these materials as catalysts with high specific surface make them very interesting subject of study. Also compressed expanded graphite composite membranes prepared from furfuryl alcohol polymers are promising for gas separation.

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