Gabriela Seydlová

• Faculty of Science, Charles University in Prague, Czech Republic

Finding effective antibiotics against multi-resistant strains of bacteria has been a challenging race. Linker-Evolved-Group-Optimized-Lipophosphonoxins (LEGO-LPPOs) are small modular synthetic antibacterial compounds targeting the cytoplasmic membrane. Here we focused on understanding the reasons for the variable efficacy of selected LEGO-LPPOs (LE...

Linker-Evolved-Group-Optimized-Lipophosphonoxins (LEGO-LPPO) are small synthetic modular peptidomimetics with promising antimicrobial activity. The LEGO-LPPO mechanism of antibacterial action has been determined to be the depolarization and disruption of bacterial membranes. Their modular nature is advantageous for fine tuning their biological prop...

Finding effective antibiotics against multi-resistant strains of bacteria has been a challenging race. LEGO-lipophosphonoxins (LEGO-LPPOs) are small modular synthetic antibacterial compounds targeting the cytoplasmic membrane. Here we focused on understanding the reasons for the variable efficacy of selected LEGO-LPPOs (LEGO-1, LEGO-2, LEGO-3, and...

In the past few decades, society has faced rapid development and spreading of antimicrobial resistance due to antibiotic misuse and overuse and the immense adaptability of bacteria. Difficulties in obtaining effective antimicrobial molecules from natural sources challenged scientists to develop synthetic molecules with antimicrobial effect. We deve...

Phage tail-like bacteriocins are now the subject of research as potent antibacterial agents due to their narrow host specificity and single-hit mode of action. In this work, we focused on the structure and mode of action of fonticins.

The alarming rise of bacterial antibiotic resistance requires the development of new compounds. Such compounds, lipophosphonoxins (LPPOs), were previously reported to be active against numerous bacterial species, but serum albumins abolished their activity. Here we describe the synthesis and evaluation of novel antibacterial compounds termed LEGO-L...

Microbial cyclic lipopeptides are an important class of antifungal compounds with applications in pharmacology and biotechnology. However, the cytotoxicity of many cyclic lipopeptides limits their potential as antifungal drugs. Here we present a structure-activity relationship study on the puwainaphycin/minutissamide (PUW/MIN) family of cyclic lipo...

Lipophosphonoxins (LPPOs) are small modular synthetic antibacterial compounds that target the cytoplasmic membrane. First-generation LPPOs (LPPO I) exhibit an antimicrobial activity against Gram-positive bacteria; however they do not exhibit any activity against Gram-negatives. Second-generation LPPOs (LPPO II) also exhibit broadened activity again...

Surfactin, a cyclic lipoheptapeptide produced by Bacillus subtilis, is a surface-active antimicrobial that targets the barrier function of lipid membranes. It inserts itself into the membrane, where it forms conductive pores. Depending on its concentration, it eventually disintegrates the membrane in a detergent-like manner. The molecular details o...

Colicin U is a protein produced by the bacterium Shigella boydii (serovars 1 and 8). It exerts antibacterial activity against strains of the enterobacterial genera Shigella and Escherichia. Here we report that colicin U forms voltage-dependent pores in planar lipid membranes; its single-pore conductance was found to be about 22 pS in 1M KCl, at pH...

The adenylate cyclase toxin-hemolysin (CyaA, ACT or AC-Hly) of pathogenic Bordetellae delivers its adenylyl cyclase (AC) enzyme domain into the cytosol of host cells and catalyzes uncontrolled conversion of cellular ATP to cAMP. In parallel, the toxin forms small cation-selective pores that permeabilize target cell membrane and account for the hemo...

Daptomycin is a calcium-dependent lipodepsipeptide antibiotic clinically used to treat serious infections caused by Gram-positive pathogens. Its precise mode of action is somewhat controversial; the biggest issue is daptomycin pore formation, which we directly investigated here. We first performed a screening experiment using propidium iodide (PI)...

Mitochondria have evolved diverse forms across eukaryotic diversity in adaptation to anoxia. Mitosomes are the simplest and the least well-studied type of anaerobic mitochondria. Transport of proteins via TIM complexes, composed of three proteins of the Tim17 protein family (Tim17/22/23), is one of the key unifying aspects of mitochondria and mitoc...

The increase in the number of bacterial strains resistant to known antibiotics is alarming. In this study we report the synthesis of novel compounds termed Lipophosphonoxins II (LPPO II). We show that LPPO II display excellent activities against Gram positive and -negative bacteria, including pathogens and multiresistant strains. We describe their...

Changes in environmental temperature represent one of the major stresses faced by microorganisms as it affects the function of the cytoplasmic membrane. In this study we have analyzed the thermal adaptation in two closely related respiratory pathogens Bordetella pertussis and Bordetella bronchiseptica. While B. pertussis represents a pathogen stric...

The capillary electrophoresis method has been developed for the analysis of phospholipid polar headgroups. The method consists in splitting the phospholipid samples by regioselective enzyme that cleaves the bond between polar headgroup and glycerol. The presented capillary electrophoresis method operates in basic pH where all the phosphoesters are...

A rapid and selective GC-MS method was optimised for the simultaneous analysis of fatty acids, phosphoethanolamine, phosphoglycerol and phosphoserine, which comprise the polar headgroups of phospholipids. Enzymatic hydrolysis was used for sample preparation to selectively release fatty acids and prevent cleavage of the phosphoester bonds of the pol...

Surfactin, an anionic lipopeptide produced by Bacillus subtilis, is an antimicrobial that targets the cytoplasmic membrane. Nowadays it appears increasingly apparent that the mechanism of resistance against these types of antibiotics consists of target site modification. This prompted us to investigate whether the surfactin non-producing strain B....

Direct injection mass spectrometry was amended with chromatographic methods in analysis of membrane phospholipids of Bacillus subtilis mutant strain, which contains four polar headgroups only (phosphatidylglycerol, phosphatidylethanolamine, phosphatidylserine, and phosphatidic acid). Phospholipid classes were determined by thin layer chromatography...

The advantages offered by established antibiotics in the treatment of infectious diseases are endangered due to the increase in the number of antibiotic-resistant bacterial strains. This leads to a need for new antibacterial compounds. Recently, we discovered a series of compounds termed lipophosphonoxins (LPPOs) that exhibit selective cytotoxicity...

Search for a potential protein binding partner(s) of LPPOs B. subtilis cell lysate was incubated either with empty streptavidin beads (EB) or with the beads couples with biotinylated LPPO (DR5690). The proteins associating with EB and LPPO were then resolved on SDS—PAGE and stained with Commassie Blue. A labeled molecular weight ladder (M) is shown...

CF release curve fitting. Time course of liposome lysis induced by DR5026 (A). Solid lines show the fit of the functions: α(1-exp(-t/τ))n to the data. Leakage of liposomes prepared from the bacterial phospholipids exhibited a biphasic behavior—in this data set the parameters for the fit of the initial phase (f(x)) were α = 29.7, τ = 175.5 s, n = 0....

Concentration-dependent cytoplasmic membrane permeabilization induced by DR5026. The increase in fluorescence intensity reflected intercalation of PI into DNA and dsRNA that occurred after PI entry into the cell upon membrane permeabilization. (PDF)

Synthesis of DR5557, biotinylated LPPO DR5690, and fluorescently labeled LPPO DR5832. (PDF)

MD simulations. (PDF)

The induction of resistance of Enterococcus faecalis and Streptococcus agalactiae to compounds DR5047 and DR5026. Minimum inhibitory concentrations of the tested LPPOs after each induction step. The induction of resistance was carried out by repeated passages of E. faecalis and S. agalactiae strains with subinhibitory concentrations of the relevant...

Structures of potential metabolites of DR5047. In all HPLC-MS analyses only intact DR5026 was detected. In the case of DR5047 (blue rectangle) less than 5% of the compound with the exact mass of 761.42 was detected. It corresponds to a monoacetylated compound—highlighted by the green rectangle—the exact position of the acetyl group has not been det...

LogD calculation. (PDF)

Antibacterial activity of LPPOs DR5557, DR5690, and DR5823. (PDF)

MTD test in mouse model—a detailed description. (PDF)

A new, fast, selective, and reliable capillary electrophoresis method has been developed for analysis of selected phosphoesters (phosphoserine, phosphoethanolamine, phosphoglycerol) and phosphate. The method is based on separation of specific phosphate containing headgroups (phosphoesters) which are cleaved from the glycerol skeleton of a phospholi...

In this study, the influence of the size and surface termination of diamond nanoparticles (DNPs) on their antibacterial activity against Escherichia coli and Bacillus subtilis was assessed. The average size and distribution of DNPs were determined by dynamic light scattering and X-ray diffraction techniques. The chemical composition of the DNPs stu...

Surfactin is a cyclic lipopeptide antibiotic that disturbs the integrity of the cytoplasmic membrane. In this study, the role of membrane lipids in the adaptation and possible surfactin tolerance of the surfactin producer B. subtilis ATCC 21332 was investigated. During a 1-day cultivation, the phospholipids of the cell membrane were analyzed at the...

In this study, we investigated the potential antibacterial properties of nanocrystalline diamond. In particular, we tested the effect of diamond nanoparticles (DNPs) on growth of the model gram-negative bacterium Escherichia coli on solid, nutrient-rich growth medium. We found that the presence of DNPs on agar plates significantly reduced the colon...

To find out membrane tolerance strategy to ethanol in Bacillus subtilis that possesses a powerful system of protection against environmental stresses. Cytoplasmic membranes of B. subtilis were severely affected by even short-term exposure to 3% (v/v) ethanol: the growth rate and membrane protein synthesis were markedly reduced, and no adaptive alte...

Surfactin, a highly powerful biosurfactant produced by various strains of the genus Bacillus, exhibits antibacterial, antiviral, antitumor and hemolytic action. This anionic cyclic lipopeptide is constituted by a heptapeptide interlinked with a β-hydroxy fatty acid. Due to its amhipathic nature surfactin incorporates into the phospholipid bilayer a...

Processes occurring in the cytoplasmic membrane of the surfactin producer Bacillus subtilis were examined during a 3-d cultivation. The fatty acid composition was found to be almost stable within this interval, except for the early stationary phase when the nonbranched, mostly C16:0 and C18:0 (high melting fatty acids), prevailed transiently in the...