Anna Poladyan

In Escherichia coli, the FhlA acts as a transcriptional activator for fdhF , hyc and hyp operons, whose gene products constitute the formate hydrogen lyase (FHL) complexes. These complexes contribute to bioenergetic regulation by consuming intracellular protons during the conversion of formate to hydrogen gas (H ₂ ). The current study elucidates the role of the FHL complex (using fhlA mutant) in the proton transport and energy transduction during the fermentation of glucose, glycerol and formate at pH 5.5. It was shown that F _O F ₁ -ATPase activity and proton flux rate were decreased in fhlA at 20 h and 72 h grown cells, compared to WT indicating the functional interplay between FHL and F _O F ₁ and highlighting its importance in proton transfer at acidic conditions. Increased number of membrane –SH groups and decreased proton conductance (C _M H ⁺ ) value in both WT and fhlA mutant were detected at 72 h, compared to 20 h, suggesting efficient energy transduction at acidic pH when H ₂ was not generated. The value of membrane potential (ΔΨ) remained unchanged in FhlA-lacking cells and was independent of growth time. Thus, bacteria regulate proton motive force (Δp) managing bioenergetic association between proton ATPase activity, FHL function and transmembrane proton gradient (ΔpH) regulation. Overall, these findings demonstrate that the FHL complex plays an essential role in coordinating proton flux, regulation of proton motive force and energy transduction in E. coli under acidic fermentative conditions through functional interplay with the F _O F ₁ -ATPase.

Vibration pathologies are one of the pressing issues in medicine and biochemistry. It is known that under the influence of vibration, the content of amide groups of proteins in tissues increases, which is one of the biochemical indicators of the organism. It has been shown that amidation occurs to a lesser extent in brain proteins of old rats exposed to vibration than in young rats. When Trifolium pratense L. flowers are added to the animals' diet, the level of amidation during vibration in old rats also decreases to a lesser extent than in young rats. However, in both cases, the level of amidation occurring is reduced by half under the influence of the plant. Based on these results, the use of T. pratense plant material is proposed as a means of reducing the negative impact of vibration on the body.

Cupriavidus necator H16 expresses O2-tolerant [NiFe]-hydrogenases (Hyds), which present promising anodic biocatalysts in enzymatic fuel cells (EFC). Bacteria also produce polyhydroxyalkanates (PHA). Depolymerization of this polymer leads to the production of crotonic acid (CA), which was estimated by the production of CA. Cheese and curd whey (SW and AW) with a combination of glycerol (G) assessed for the growth of C. necator H16, PHA accumulation, and Hyds activity. Bacterial growth, PHA accumulation, and Hyd activity were observed under microaerophilic conditions for 7 days. Prolonged bacterial growth was observed in glycerol-supplemented samples. Maximum biomass formation was observed on the 5th day in AW (OD6003.1) and on the 6th day in SW+G (OD600 3.3). pH and ORP decrease and Hyd activity were detected under all conditions; however, SW+G sample showed enhanced Hyd activity (0.17±0.03 U·min−1·g−1CDW) compared with G-free samples. A clear correlation was shown between PHA accumulation and Hyd activity: with the Hyd activity decline, CA concentration increased, reaching up to 2.4 ± 0.05 mg/mL. The results will advance hydrogen-based EFC systems as well as the sustainable synthesis of biodegradable PHAs.

Cupriavidus necator H16 possesses oxygen-tolerant [NiFe] hydrogenases, which can oxidize H2 in the presence of oxygen (O2), easily adapts between autotrophic and heterotrophic lifestyles, and has significant biotechnological potential. Aim. Тo investigate the growth characteristics of C. necator H16 using different nutrient media and heavy metal, particularly iron ions, for determination of optimal conditions maximizing Hyd activity. Methods. C. necator H16 was cultivated heterotrophically using Nutrient Broth (NB) and Fructose-Nitrogen (FN) minimal growth media, pH 7.0. The effects of iron (Fe(III)) ions on the activity of the bacterial H2-oxidizing Hyds (hydrogenases) and its growth properties were investigated. Bacterial growth was monitored by determining the optical density, OD570, under 570 nm. The total H2-oxidizing Hyd activity of whole cells of C. necator H16 was measured over a 72-hour growth period. Results. In the control samples without adding Fe ions, the activity was observed in the range of 0.5–0.2 U mg–1 cell dry weight (CDW) in both NB and FN media for up to 48 hours of bacterial growth. However, when 54 µM Fe ions were added to the media, the H2-oxidizing Hyd activity increased approximately 2–3 fold after 24 and 48 hours in both FN and NB media, compared to the control samples. Additionally, in the samples supplemented with Fe ions, a shift in the ORP from positive to negative values was observed, primarily starting from the exponential growth phase. After 24 hours, the NB medium showed stimulated biomass formation and Hyd activity compared to the FN medium. Conclusions. These findings suggest that certain conditions, such as the presence of higher amounts of iron ions and specific growth media, can enhance the Hyd activity during bacterial heterotrophic growth.

Fruit peels as a lignocellulosic biomass are rich in sugars and nutrients, thus making them favorable medium for yeast growth and various materials production. This study showed the potential of fuit peels hydrolysate (FPH) as a substrate for single-cell protein (SCP) and bioethanol production, depending on substrate concentration, pH, oxygen availability. The highest specific growth rate (0.50 ± 0.01 h^- 1) was observed at pH 6.5 in non-diluted hydrolysates, which corresponds to ~ 250.0 g L^- 1 carbohydrates and 0.2 g L^- 1 total nitrogen concentration. The maximal protein production yield (~ 50% by dry weight) was recorded during 24 h cultivation of S. cerevisiae ATCC 13,007 under oxygen-limited conditions at pH 6.5, while the maximal ethanol production (109.00 ± 4.00 g L^- 1) was observed at 6 h for S. cerevisiae ATCC 9804 and ATCC 13,007 starins under aerobic conditions. The maximal fermentation efficiency (~ 99%) was observed at 24 h cultivation of S. cerevisiae ATCC 13,007 in 2-fold diluted hydrolysate under oxygen-limited conditions at pH 6.5, while the same strain exhibited the highest carbon conversion efficiency (CCE) (~ 56%) during aerobic cultivation in 2-fold diluted hydrolysate at pH 3.0. Thus, FPH can serve as a favorable substrate for SCP and bioethanol production using S. cerevisiae strains ATCC 13,007 and ATCC 9804. Nitrogen supplementation and combination with fruit and vegetable processing industries could further enhance SCP production and process efficiency which aligns with circular economic strategy.

Hydrogenases (Hyds) are microbial enzymes that reversibly catalyze the oxidation of molecular hydrogen (H₂), playing a critical role in biohydrogen metabolism [1-3]. These include oxygen-resistant biological [NiFe]-hydrogenases, which have attracted great interest for their application in hydrogen fuel cell (H₂-FC) technologies. The results provide insight into the potential of bioelectrochemical-based systems for sustainable energy production. The bioelectrocatalytic efficiency of E. coli bacteria immobilized on the electrode surface in a volume of 3 μl (1.5 mg cell dry weight) was studied under the conditions of 0.2% glucose fermentation in peptone medium at pH-7.5 [4]. In this study, the electrochemical measurements were performed using a two-electrode system equipped with a computer potentiostat, specifically a hydrogen fuel cell voltammetry (HFCV). The wild-type E. coli BW25113, the septuple (BW25113hyaB hybC hycA fdoG ldhA frdC aceE) the F0F1-ATPase-defective, and the Hyd defective hyaB, hybC, hycE, hyfG mutant strains were used in the experiments. Maximal catalytic activity was observed in the hyaB and hyfG mutants, being stimulated ∼2-fold and ∼1.6-fold compared to the wild type, reaching values of ∼1.26 ± 0.02 V and ∼0.98 ± 0.02 V, respectively. The effect of the 10 mM N,N′-dicyclohexylcarbodiimide (DCCD), the F0F1-ATPase inhibitor, on the catalytic activity of Hyd enzymes was observed. It was shown that for all strains, the reading of the voltammeter decreased ~1.5 times, reaching the readings recorded by the F0F1-ATPase-defective strain. Interestingly, in the case of the the septuple mutant strain, DCCD recorded a stimulating rather than a suppressive effect. The results obtained indicate the great potential of bacteria as anodic biocatalysts and demonstrate the need for further studies.

Contemporary higher education demands innovative teaching approaches that integrate scientific thinking, collaborative skills, and the development of creative solutions. The aim of this study is to develop and examine the LEGO Serious Play (LSP) method as an innovative approach in the teaching of university-level courses. Specifically, the application of this method was explored within the course "From Research to Production: Perspectives and Practice" offered in the Master's program in Applied Microbiology at Yerevan State University (YSU). The method was designed to foster systems and project-based thinking through play, with the goal of bridging the gap between scientific research outcomes and their application in industrial processes. As a result, students used LEGO sets to physically model their ideas, presented their projects through narrative storytelling, worked in a team-based environment, and participated in the co-creation of solutions. The outcomes of applying the method indicate an increase in student engagement, as well as enhancement of their creative and collaborative skills. The LSP method can serve as an effective tool to enhance learning processes and stimulate analytical and applied thinking in biomedical, natural sciences, and other educational programs.