Biology subjects

Vasilchenko, A. S.

Publications and source records attributed to Vasilchenko, A. S..

4 recordsLinked to original sources

bioRxiv2024

Macrolactin A Is an Inhibitor of Protein Biosynthesis in Bacteria

The macrolide antibiotic, macrolactin A (McA), has been known for its antimicrobial properties since the late 1980s, but the mechanism of its antibacterial activity is still unknown. In this study, we investigated the microbiological and molecular characteristics of McA antimicrobial activity. McA effect on bacteria was found to be both bacteriostatic and bactericidal, depending on species and strains. Regarding the mechanism of action of McA, the following important results were obtained: 1) using in vivo and in vitro systems, we showed that McA is an inhibitor of protein synthesis in bacteria; 2) the concentration of McA required to inhibit protein synthesis in the E. coli cell-free model was found to be 50 times lower than the concentration required in the S. aureus cell-free model; 3) the toe-printing assay revealed that McA inhibits the first step of elongation stage of protein synthesis; 4) we identified single and multiple nucleotide polymorphisms in the gene encoding the translation elongation factor Tu (EF-Tu) by annotating the genomes of McA-resistant Bacillus pumilus McAR and its parental strain. Molecular modeling showed that the McA molecule can form non-covalent bonds with amino acids at the interface of domains 1 and 2 of EF-Tu, characterized by a relatively high docking score. Overall, our study demonstrated that McA acts as an elfamycin-like antibiotic (targeting EF-Tu), addressing a substantial gap in our understanding of the mechanism of action of macrolactin A, a representative member of macrolides.

microbiology↗

bioRxiv2022

Antibacterial Action of Gliotoxin Realized Through Interaction with Tiol-containing Proteins

Gliotoxin is a secondary metabolite of various fungi belonging to the class of epipolythiodioxopiperazines. The mechanism of gliotoxin cytotoxic activity on eukaryotes is established, while the precise interaction between gliotoxin and bacteria has not been clarified yet. The aim of this study was evaluating the gliotoxin action on model Gram-positive and Gram-negative bacteria. It was found that gliotoxin uptake rate was higher in S. epidermidis than E.coli. However, for Gram-negative bacteria, the bactericidal effect is achieved at higher doses of gliotoxin (10-100 g/ml), compared to Gram-positive ones (0.75-6.25 g/ml). Bactericidal effect had developed in 4 hours, which was the same for both E. coli MG1655 and S. epidermidis. Novel chip-based bioluminescent sensor with gel-immobilized E.coli MG1655 pKatG-lux revealed oxidative stress. However, pre-incubation of bacteria with Trolox did not affect the bactericidal properties of gliotoxin, while 2-merkaptoethanol and reduced glutathione significantly reduced gliotoxins bactericidal property. Another bioluminescent sensor E.coli MG1655 pIbpA-lux revealed heat shock stress in bacteria treated with gliotoxin. At the cellular level, exposure bacteria with gliotoxin accompanied by disruption of bacterial membranes.

microbiology↗

bioRxiv2022

Generation of Gausemycin A-resistant Staphylococcus aureus

Gausemycins A and B are the first members of the novel lipoglycopeptides family produced by Streptomyces roseoflavus INA-Ac-5812, which showed the ability to fight clinically important Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus. However, new antibiotics need to be studied in depth to determine their full potential. In this study, we concentrated our efforts to investigate resistance emerging within S. aureus upon gausemycin A application. Using serial passaging of S. aureus FDA209P in increasing concentrations of gausemycin A, we obtained the resistant variant S. aureus 5812R which are 80-times more resistant comparing to the origin strain. Moreover, obtained resistance is stable, since 15 passages in a drug-free medium did not restore bacterial susceptibility to gausemycin A. Elucidating of the differences between resistant and parent strains was concerned antibiotic cross-resistance, structure of bacterial membrane, and response at genetic level. Susceptibility testing of S. aureus 5812R revealed the acquisition of cross-resistance to daptomycin, cefazolin, and tetracycline, while resistance to vancomycin, nisin and ramoplanin absence. The composition of fatty acids constituting the cytoplasmic membrane of S. aureus 5812R, was represented by increased content of anteiso- branched chain fatty acids, while iso-branched chain fatty acids was decreased comparing the origin S. aureus FDA209P strain. The relative expression of the cls gene catalyzing the synthesis of cardiolipin in the resistant cells was higher compared to the S. aureus FDA209P.

microbiology↗

bioRxiv2022

2,4-Diacetylphloroglucinol Against Candida albicans: Biofilm Formation, Aspartyl Protease Production and Ultrastructure Changes

The dimorphic fungus Candida albicans is the one of the most important opportunistic pathogen of human. The use of fungicides against candida encounters sub-inhibitory effects, which are fungal stress response and undesirable for the host. In this work we investigated antifungal action of 2,4-diacetylphloroglucinol against Candida albicans ATCC 10231 with focus on they biofilm forming ability under the treatment. It was found that 2,4-DAPG reduced ability of candida cells to form biofilm, but complete inhibition or irradiation did not achieve. Moreover, Candida albicans cells in the adherent state have been characterized by reduced susceptibility to 2,4-DAPG comparing to planktonic cells. Investigation of mechanisms which could be explain the antibiofilm action of 2,4-DAPG has revealed the reduction in cells surface hydrophobicity and inhibition of yeast-to-hyphae transition. Biofilms formed under the sub-inhibitory concentrations of 2,4-DAPG were depleted in protein and carbohydrate content. Furthermore, we microscopically visualized the treated biofilms and have revealed numerous cannels localized on hyphae and associated with secretion of aspartyl proteases (Sap). We assumed what excretion of Sap is triggered by reactive oxygen generated upon affecting of mitochondrion respiration by 2,4-DAPG. Introducing of antioxidant Trolox simultaneously with 2,4-DAPG lead to reducing of Sap production. Thus, production of aspartyl proteases is a one of undesirable side effect of Candida albicans treatment, but using 2,4-DAPG in combination with antioxidants is the solution to overcome it.

microbiology↗