Biology subjects

Dejmek, M.

Publications and source records attributed to Dejmek, M..

5 recordsLinked to original sources

bioRxiv2026

Characterization of the N7 RNA cap methyltransferase from Trichomonas vaginalis and inhibitor discovery

RNA cap formation is essential for eukaryotic gene expression, yet the enzymes responsible for cap methylation remain poorly characterized in many eukaryotic parasites, including Trichomonas vaginalis. Here, we functionally and structurally characterized the RNA guanine-N7 methyltransferases (MTases) from T. vaginalis, TvRNMT1 and TvRNMT2. We determined the crystal structure of TvRNMT1 in complex with S-adenosylhomocysteine (SAH) at 1.6 A resolution and additional structures with the inhibitors sinefungin and OBO101, defining the architecture and ligand-recognition properties of its active site. Screening of SAH analogues identified several inhibitors with submicromolar to low-micromolar activity against both homologues. Molecular docking and modelling of capped RNA binding further delineated the inhibitor- and RNA-binding regions of TvRNMT1 and identified residues that may contribute to cap recognition. These results define the molecular basis of ligand recognition by T. vaginalis RNA cap MTases and establish a structural framework for targeting parasite RNA capping.

molecular biology↗

bioRxiv2026

Branched C7-Substituted 7-Deaza-SAH Analogues Occupy the Entire SAM-Binding Pocket of Mpox Virus VP39 and Dengue Virus NS5 Methyltransferases

Viral RNA-cap MTases are attractive targets for antiviral drug development. We previously identified C7-substituted 7-deaza-SAH analogues as potent inhibitors of the mpox virus 2'-O-MTase VP39. Here, we used structure-guided design to develop branched C7-substituted analogues intended to engage multiple hydrophobic regions of the VP39 SAM-binding pocket. The synthesized compounds were characterized using biochemical and crystallographic approaches. Several analogues effectively inhibited VP39, with the most potent compound displaying an IC50 in the tens-of-nanomolar range. Crystal structures of VP39 in complex with STM1187 and STM1189 confirmed that the branched aromatic substituents extend towards hydrophobic regions adjacent to the SAM binding site. The precise ligand conformations were strongly influenced by linker geometry and the branching groups mode of attachment. STM1078 also inhibited DENV3 NS5 MTase with submicromolar potency, and the complexs structure revealed a conserved binding mode of the SAH-like core accompanied by conformational adaptability of the branched substituent. These results demonstrate how three-dimensional expansion from the 7-deaza position can generate potent inhibitors capable of binding structurally distinct viral MTases.

molecular biology↗

bioRxiv2025

Bemnifosbuvir and remdesivir inhibit tick-borne encephalitis virus infection in complementary in vitro and ex vivo disease models

The geographical distribution and incidence of tick-borne encephalitis (TBE) have steeply increased over the past decades, raising to represent a major health concern in Asia and Europe. Symptoms of TBE, caused by infections with tick-borne encephalitis virus (TBEV), range from mild, flu-like symptoms to severe neurological disease, often accompanied by long-term sequelae persisting for several years following pathogen encounter. While effective vaccines against TBEV are available, no antiviral drugs are currently approved and therapeutic options for patients suffering from TBE are limited to supportive measures. Compounds able to disrupt viral nucleic acid synthesis bear the potential of effectively limiting viral replication and spread. Seeking to fill the therapeutic gap, we evaluated the efficacy of a panel of approved and investigational antiviral compounds in containing TBEV infection. Combining several cell lines, human neural organoids and organotypic rat brain slice cultures, we found that the nucleoside analogs remdesivir and bemnifosbuvir efficiently limit viral replication. Through infectious virus quantification, immunofluorescence analysis and flow cytometry, we report significant, dose-dependent reduction of viral loads across all models used, with inhibition observed at low doses for both drugs. Notably, while we observed bemnifosbuvir to be well tolerated, we report important cytotoxicity of remdesivir when applied to human neural organoids. Our findings identify bemnifosbuvir and remdesivir as novel treatment strategies for TBE, providing an accessible and timely response to a clinical challenge of pressing concern.

microbiology↗

bioRxiv2024

Mechanistic Insights into Alcohol-Induced DNA Crosslink Repair by the Fanconi Anemia Nuclease Slx4-Xpf-Ercc1

During cell division, DNA replication stalls upon alcohol-derived interstrand crosslink, employing DNA repair pathways. Acetaldehyde, a toxic metabolite of alcohol, can induce DNA crosslinks between complementary DNA strands, causing significant genomic instability. The repair of acetaldehyde DNA crosslinks (AA-ICLs) is vital for maintaining genome integrity and preventing mutagenic events. The Fanconi anemia (FA) repair pathways involvement in fixing the AA-ICLs is crucial, ensuring cellular homeostasis and safeguarding genomic stability. Dysregulation of the FA pathway has been linked to Fanconi anemia, a rare genetic disorder characterized by hypersensitivity to DNA-damaging agents, including acetaldehyde. However, the precise mechanism of the repair and incision of AA-ICLs is unknown. Here, we demonstrate the role of the FA pathway nuclease SLX4-XPF-ERCC1 (SXE) in the repair of AA-ICL. We have generated site-specific AA-ICL within the replication fork, and we found SXE to excise the crosslink from the replication fork, bringing evidence of its role in the repair of alcohol-induced DNA lesions. This nuclease complex performs two precise incisions around the lesion. We also demonstrate that the SXE nuclease excises an abasic site interstrand crosslink in a similar manner. Given the evidence in conjunction with previous repair studies that have been conducted, our work suggests that SXE is a versatile nuclease complex.

biochemistry↗

bioRxiv2023

Fluorinated cGAMP analogs, which act as STING agonists and are not cleavable by poxins: structural basis of their function

The Stimulator of Interferon Genes (STING) plays a crucial role in the cGAS-STING pathway of innate immunity, detecting DNA in the cytoplasm and defending against certain cancers, viruses, and bacteria. We designed and synthesized fluorinated carbocyclic cGAMP analogs, MD1203 and MD1202D (MDs), to enhance their stability against nucleases and their affinity for STING. These compounds demonstrated exceptional activity against wild-type STING and all its allelic variations, including the hard-to-target REF isoform. Despite their distinct chemical modifications relative to the canonical CDNs, such as the substitution of guanine with hypoxanthine and the fluorination of the (pseudo)ribose ring, crystallographic analysis revealed a consistent binding mode with STING. Importantly, these compounds were resistant to cleavage by viral poxin nucleases. The crystallographic analysis of poxin/MD complexes unveiled their binding mode at the interface of poxin monomers, with dynamic adenine base orientations. Interestingly, MDs-bound poxin adopted an unliganded-like conformation, distinct from the conformation of cGAMP-bound poxin. Moreover, when MDs were in complex with poxin, they exhibited a different conformation than cGAMP when bound to poxin; in fact, it closely resembled the conformation observed when MDs were bound to STING. In conclusion, the development of MD1203 and MD1202D, showcases their potential as potent STING activators with remarkable stability against poxin-mediated degradation--a crucial characteristic for future development of antivirals.

molecular biology↗