Institute of Biomedical Chemistry

The search and identification of blood serum proteins with high osmotic activity (osmotically active proteins, OAP) were carried out in albumin-containing fish, northern Esox lucius L., using electrophoresis and MALDI mass spectrometry. Several proteins with high negative charge (8 extracellular and 1 intracellular) were found and identified in the pike blood serum. The total relative content (TRC) of all OAP was ~60%. The relative content (RC) of individual OAP—hemopexin, proteinase inhibitors, apopoprotein A-I (in high density lipoproteins) and albumin—were ~30, ~10, ~12, and 3.6%, respectively; intracellular protein Grb14 was represented by trace amounts. According to the annotations of Gene Ontology, the main functions of OAP are related to the protection and transport. At the same time, the high negative charge of OAP suggests their high osmotic activity. The comparison of OAP lists in albumin-containing pike and albumin-free fishes shows that they are consistent across all extracellular OAP except for albumin, which is negligible in the pike serum. In light of the albumin-free model of capillary exchange, these facts do not point to a key role of albumin in osmotic homeostasis control in fish. The multiplicity of OAP in the blood of teleost fishes distinguishes them from mammals, where serum albumin is a key factor controlling capillary fluid exchange.

ABSTRACT Tuberculosis persists as a major global health threat, significantly exacerbated by the rise of drug-resistant strains. Cytochrome P450 of 124 family CYP124 from Mycobacterium tuberculosis (CYP124), implicated in host sterol metabolism and bacterial virulence, represents an emerging and promising therapeutic target. While its precise physiological role was previously debated, CYP124’s confirmed ability to metabolize immunomodulatory host sterols underscores its pharmacological relevance. Utilizing surface plasmon resonance binding assays and UV-Vis spectral titration screening, we identified nine novel non-azole ligands for CYP124 from a library of 32 plant-derived and marine natural compounds. Among these hits, (25S)-5α-cholestane-3β,4β,6α,7α,8,15β,16β,26-octaol (termed 15β-octaol) and henricioside H 2 (HD-4) induced characteristic difference spectra and formed long-lived inhibitory complexes with CYP124, exhibiting dissociation half-lives of 181 min and 65 min, respectively. However, their inhibitory potency was moderate, with IC 50 values of approximately 86 μM for 15β-octaol and exceeding 100 μM for HD-4. Complementary in silico molecular docking and analysis identified key conserved hydrophobic residues within the CYP124 active site crucial for ligand binding, suggesting a shared pharmacophore. Furthermore, structural similarity analysis revealed that 37 human endogenous metabolites, including known immunoregulatory sterols, bear resemblance to the identified CYP124 ligands. This finding points towards a potential sterol-mediated interplay at the host-pathogen interface. Collectively, these results provide a foundation for the future development of mechanism-based CYP124 inhibitors as therapeutics against multidrug-resistant tuberculosis.

Background: Human cell lines underpin modern biomedical research, offering reproducibility, standardisation, and unrestricted access to biological material. Among the 1206 human lines documented in the Human Protein Atlas, in vitro systems overcome the ethical and technical constraints of primary tissues. The liver is an organ of intricate structure, diverse physiological roles, and limited in vitro viability. Liver-derived cell lines are increasingly used to address the growing burden of liver disease and to accelerate pharmaceutical development, yet their capacity to replicate native hepatic functions remains uncertain. The mutational profiles and expression patterns of hepatocyte-characteristic genes provide critical benchmarks for their suitability for pharmacology and toxicology. Methods: Here, we systematically compare ten widely used hepatic cell lines (HepG2, Huh7, Hep3B, LX-2, HepaRG, HLF, HLE, MHCC97H, SK-Hep1, PLC/PRF/5) with primary hepatocytes and liver tissue, focusing on drug-metabolizing enzyme (DME) gene expression. Beyond literature synthesis, we analysed pre-processed RNA-seq expression data. Results: Overall, among the models examined, the HepaRG cell line shows the greatest similarity to liver and primary hepatocytes, most faithfully reproducing the expression patterns of DME genes. HepG2, Hep3B, and Huh7 form a cluster that retains only a subset of hepatic characteristics. Other models display more pronounced deviations from the reference profile and are generally used for specialized applications. Thus, no universal cell line exists that can fully substitute for the liver. Each model has its own limitations and biases in the expression profile of DME genes, which must be carefully considered when selecting an appropriate system for specific research objectives.

Insufficient fruit intake is a major contributor to the development of non-communicable diseases, as the global average of daily fruit consumption remains far below the recommended levels. Apples are among the most widely consumed fruits worldwide, making them an ideal target for nutritional enhancement. Enhancing the content of health-promoting compounds within apples offers a practical way to increase bioactive intake without requiring major dietary changes. This review evaluates which of the 41 biologically active compounds considered in this article can reach physiologically relevant intake levels at the current average daily consumption of cultivated and wild apples. Comparative analysis shows that wild apples consistently contain higher concentrations of phenolic compounds and organic acids than cultivated varieties, in some cases by more than tenfold. At the average daily fruit intake of 121.8 g, wild species provide effective doses of epicatechins, anthocyanins, chlorogenic acid, and malic acid. In contrast, cultivated apples reach this level only for chlorogenic acid. Notably, less than 50 g of wild apple is sufficient to supply physiologically relevant amounts of several polyphenols. These findings highlight the potential of wild apple species as donors of bioactive compounds and provide a framework for breeding future apple cultivars that combine consumer appeal with enhanced health benefits.

The activity of cytochrome P450 enzymes decreases in older adults, which can lead to toxic effects from polypharmacy. Cytochromes P450 are the most significant enzymes involved in the metabolism of foreign compounds, including pharmaceutical substances. Vitamin B2, or riboflavin (RF), is a potent antioxidant that is vital for the body and participates in numerous enzyme-catalyzed redox reactions. RF is phosphorylated intracellularly to form flavin mononucleotide (FMN), which is further metabolized into flavin adenine dinucleotide (FAD). The active site of the NADPH-dependent cytochrome P450 reductase (CPR), a redox partner of CYP enzymes, is necessary for the catalytic functions of cytochromes P450. The active site of reductase is a complex formed by two types of vitamin B2, such as flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN). In our study, we investigated the impact of the phosphorylated form of vitamin B2, FAD, and FMN on the catalytic activity of cytochrome P450 2C9 (CYP2C9) towards non-steroidal anti-inflammatory medications diclofenac and naproxen. It was shown that FAD significantly enhanced the catalytic efficiency of CYP2C9. The 4-hydroxylation of diclofenac was enhanced by 148 ± 10%. The O-demethylation of naproxen showed an increase of 120 ± 14%. Based on these data, we can assume that intake of vitamin B2 (riboflavin) improves catalytic efficiency of CYP2C9. This finding is essential for the modulation of catalytic activity of CYP2C9. The proposed electroanalytic approach is a sensitive and robust method for drug metabolism assay.

RNA-based drugs hold significant potential, offering promising new treatments for a wide range of diseases, especially those with a genetic basis. By leveraging RNA interference (RNAi) and other RNA-mediated mechanisms, these therapies can precisely modulate gene expression and address the root causes of genetic defects. RNA-based drugs hold significant potential for treating a range of diseases. However, the transition of these therapies from laboratory research to clinical applications has encountered hurdles. This review explores the composition and outcomes of clinical trials for various modified short RNA drugs. We detail their mechanisms of action, delivery systems—with a focus on lipid nanoparticles and N-acetylgalactosamine (GalNAc) conjugates—and clinical efficacy in treating conditions such as transthyretin (TTR) amyloidosis. Our analysis reveals that while several RNAi-based drugs have achieved clinical approval, a critical unmet need remains: advanced delivery systems capable of precisely targeting diverse tissues, particularly outside the liver. We also underscore the importance of rigorous target validation utilising sophisticated bioinformatics tools and in vitro/in vivo assays to minimise off-target effects and ensure robust therapeutic efficacy. This review proposes a novel framework for optimising RNA drug development, emphasising the crucial interplay between delivery strategies, target specificity, and understanding disease-specific target biology.

This paper summarizes the results of multi-year studies performed by our research team, focusing on an analysis of protein recoding mediated by messenger RNA editing by ADAR adenosine deaminases. Searching for ADAR-mediated protein recoding was performed in the central nervous system of the model organisms, fruit fly and mouse, as well as in the human proteomic datasets. The proteogenomic approach has made it possible to identify dozens of editing events in the proteome, thus validating the results of transcriptomic studies. The observed recoding events in animals, ranging from insects to mammals, mainly affect the cytoskeletal components and proteins involved in synaptic transmission. In humans, recoding changes are most often observed in the central nervous system or tumor tissues. Over 15 million editing sites have been identified in humans; only a few thousand of those can potentially yield amino acid substitutions. Using a proteogenomic approach, dozens of protein recoding sites are identified, demonstrating their origin in ADAR RNA editing. Moreover, this revealed that the level of recoding at specific sites is not directly related to the abundance of ADAR enzymes per se or their target proteins. The recoding processes probably have differential regulation of interactions at the mRNA level that is yet to be clarified.

One of the avenues for the development of functional gradient additive manufacturing is the creation of four-dimensional (4D) printed structures for soft robotic gripping, achieved by combining fused deposition modeling 3D printing with soft hydrogel actuators. This work proposes a conceptual approach to creating an energy-independent soft robotic gripper, consisting of a modified 3D printed holder substrate made from thermoplastic polyurethane (TPU) and an actuator based on a gelatin hydrogel, allowing programmed hygroscopic deformation without using complex mechanical constructions. The use of a 20% gelatin-based hydrogel imparts soft robotic biomimetic functionality to the structure and is responsible for the intelligent stimulus-responsive mechanical functionality of the printed object by responding to swelling processes in liquid environments. The targeted surface functionalization of thermoplastic polyurethane in an argon–oxygen environment for 90 s, at a power of 100 W and a pressure of 26.7 Pa, facilitates changes in its microrelief, thus improving the adhesion and stability of the swollen gelatin on its surface. In this work, the morphological (scanning electron microscopy and atomic force microscopy), colloidal, chemical (FTIR), physicochemical (swelling), biological (cell biocompatibility), and performance evaluations of the TPU/gelatin gripper were studied. The realized concept of creating 4D printed biocompatible comb structures for macroscopic underwater soft robotic gripping can provide noninvasive local gripping, transport small objects, and release bioactive substances upon swelling in water. The resulting product can therefore be used as a self-powered biomimetic actuator, an encapsulation system, or soft robotics.