Biology subjects

Loginov, D.

Publications and source records attributed to Loginov, D..

3 recordsLinked to original sources

bioRxiv2025

Nanobodies as therapies for loss-of-function misfolding diseases.

Misfolding diseases that result in loss of function represent a considerable burden for both individuals and society. Primary hyperoxaluria type 1 (PH1) is a rare genetic disorder caused by mutations in the alanine:glyoxylate aminotransferase 1 (AGT) enzyme. The underlying molecular mechanisms causing PH1 are associated with protein misfolding (enhanced aggregation and mitochondrial mistargeting). The main therapeutic approach to increase patients lifespan and quality of life is a double kidney and liver transplantation. Alternative treatments such as gene and enzyme replacement and pharmacological chaperones are currently being introduced, but other alternatives are necessary. In this work, we developed and characterized a novel biotechnological approach using six single-domain nanobodies (NB-AGT-1 to -6) as potential therapeutics for PH1 misfolding. We show that NB-AGTs are very stable proteins and bind to pathogenic and non-pathogenic variants of AGT with extreme affinities (with Kd values from low nM to low pM). Structural studies showed that NB-AGTs bind to different epitopes of AGT with selectivity for different AGT variants. Experiments in cellular PH1 models showed that internalization of engineered NB-AGT-3 enhanced the specific activity of disease-associated variants. Overall, we show that NBs are a novel and promising approach to treat PH1 and other loss-of-function misfolding diseases.

biophysics↗

bioRxiv2025

New structural insights into the control of the retinoic acid receptors RAR/RXR by DNA, ligands and transcriptional coregulators

Retinoic acid receptors (RARs) are ligand-dependent transcription factors essential for various biological processes, including embryogenesis, differentiation, and apoptosis. RARs function as heterodimers with retinoid X receptors (RXRs) and regulate gene expression via retinoic acid response elements (RAREs). Their transcriptional activity is modulated by coregulators, with corepressors maintaining repression in the absence of ligand and coactivators enabling transcription upon ligand binding. Structural studies reveal that DNA binding induces conformational changes affecting coregulator interactions. However, the precise structural organization of RAR/RXR-coregulator complexes and the allosteric influence of DNA on receptor function remain incompletely understood. Our study presents an integrative analysis of the RAR/RXR heterodimer bound to four distinct and relevant RAREs (DR0, DR1, DR5, and IR0) in complex with either a corepressor (NCoR) or a coactivator (TIF-2) nuclear receptor interaction domain. By combining small-angle X-ray scattering, hydrogen/deuterium exchange mass spectrometry and molecular dynamics simulations, we revealed that the heterodimer adopts distinct conformations depending on the DNA sequence, influencing interdomain distances and receptor interactions. Additionally, we uncovered the dynamic interplay between ligand, DNA, and coregulator binding. This study provides new insights into the structural features of coregulator proteins and highlights the allosteric influence of RAREs on receptor function.

biochemistry↗

bioRxiv2021

Shotgun proteomic profiling of dormant, 'non-culturable' Mycobacterium tuberculosis

Dormant cells of Mycobacterium tuberculosis, in addition to low metabolic activity and a high level of drug resistance, are characterized by non-culturability - a specific reversible state of the inability of the cells to grow on solid media. The biochemical characterization of this physiological state of the pathogen is only superficial, pending clarification of the metabolic processes that may exist in such cells. In this study, applying LC-MS proteomic profiling, we report the analysis of proteins accumulated in dormant, non-culturable M. tuberculosis cells in an in vitro model of self-acidification of mycobacteria in the post-stationary phase, simulating the in vivo persistence conditions. This approach revealed the accumulation of a significant number of proteins (1379) in cells after 4 months of storage in dormancy; among them, 468 proteins were significantly different from those in the actively growing cells and bore a positive fold change (FC). Differential analysis revealed the proteins of the pH-dependent regulatory system phoP and allowed the reconstruction of the reactions of central carbon/glycerol metabolism, as well as revealing the salvaged pathways of mycothiol and UMP biosynthesis, establishing the cohort of survival enzymes of dormancy. The annotated pathways mirror the adaptation of the mycobacterial metabolic machinery to life within lipid-rich macrophages, especially the involvement of the methyl citrate and glyoxylate pathways. Thus, the current in vitro model of M. tuberculosis self-acidification reflects the biochemical adaptation of these bacteria to persistence in vivo. Comparative analysis with published proteins with antigenic properties makes it possible to distinguish immunoreactive proteins (40) among the proteins bearing a positive FC in dormancy, which may include specific antigens of latent tuberculosis. Additionally, the biotransformatory enzymes (oxidoreductases and hydrolases) capable of prodrug activation and stored up in the dormant state were annotated. These findings may potentially lead to the discovery of immunodiagnostic tests for early latent tuberculosis and trigger the discovery of efficient drugs/prodrugs with potency against non-replicating, dormant populations of mycobacteria.

microbiology↗