Biology subjects

Vicente, F.

Publications and source records attributed to Vicente, F..

3 recordsLinked to original sources

bioRxiv2024

Exploring the druggability of the UEV domain of human TSG101 in search for broad-spectrum antivirals

The ubiquitin E2 variant domain of TSG101 (TSG101-UEV) plays a pivotal role in protein sorting and virus budding by recognizing PTAP motifs within ubiquitinated proteins. Disruption of TSG101-UEV/PTAP interactions has emerged as a promising strategy for the development of host-oriented broad-spectrum antivirals with low susceptibility to resistance. TSG101 is a challenging target characterized by an extended and flat binding interface, low affinity for PTAP ligands, and complex binding energetics. Here, we assess the druggability of the TSG101-UEV/PTAP binding interface by searching for drug-like inhibitors and evaluating their ability to block PTAP recognition, impair budding, and inhibit viral proliferation. A discovery workflow was established combining in vitro miniaturized HTS assays and a set of cell-based activity assays including high-content bimolecular complementation, virus-like particle release measurement, and antiviral testing in live virus infection. This approach has allowed us to identify a set of chemically diverse molecules that block TSG101-UEV/PTAP binding with IC50s in the low M range, and able to disrupt the interaction between full-length TSG101 and viral proteins in human cells and inhibit viral replication. State-of-the-art molecular docking studies reveal that the active compounds exploit binding hotspots at the PTAP binding site, unlocking the full binding potential of the TSG101-UEV binding pockets. These inhibitors represent promising hits for the development of novel broad-spectrum antivirals through targeted optimization and are also valuable tools for investigating the involvement of ESCRT in the proliferation of different virus families and study the secondary effects induced by the disruption of ESCRT/virus interactions. ImportanceMany viruses rely on the interaction between TSG101 and viral proteins containing PTAP motifs for their proliferation. Here we show that these interactions can be efficiently blocked by drug-like compounds that impair budding and replication of viruses from different families. We have also provided valuable insights into the determinants of high affinity for these small molecule inhibitors that open new avenues for developing the identified candidates into broad-spectrum antivirals with low susceptibility to resistance.

molecular biology↗

bioRxiv2023

Establishment of a screening platform based on human coronavirus OC43 for the identification of microbial natural products with antiviral activity

Human coronaviruses (HCoVs) cause respiratory tract infections and are of great importance due to the recent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. Human betacoronavirus OC43 (HCoV-OC43) is an adequate surrogate for SARS-CoV-2 because it infects the human respiratory system, presents a comparable biology, and is transmitted in a similar way. Its use is advantageous since it only requires biosafety level (BSL)-2 infrastructure which minimizes costs and biosafety associated limitations. In this report, we describe a high-throughput screening (HTS) platform to identify compounds that inhibit the propagation of HCoV-OC43. Optimization of assays based on inhibition of the cytopathic effect and virus immunodetection with a specific antibody, has provided a robust methodology for the screening of a selection of microbial natural product extracts from the Fundacion MEDINA collection. Using this approach, a subset of 1280 extracts has been explored. Of these, upon hit confirmation and early LC-MS dereplication, 10 extracts were identified that contain potential new compounds. In addition, we report on the novel antiviral activity of some previously described natural products whose presence in bioactive extracts was confirmed by LC/MS analysis. IMPORTANCEThe COVID-19 pandemic has revealed the lack of effective treatments against betacoronaviruses and the urgent need for new broad-spectrum antivirals. Natural products are a valuable source of bioactive compounds with pharmaceutical potential that may lead to the discovery of new antiviral agents. Specifically, compared to conventional synthetic molecules, microbial natural extracts possess a unique and vast chemical diversity and are amenable to large-scale production. The implementation of a high-throughput screening platform using the betacoronavirus OC43 in a human cell line infection model has provided proof of concept of the approach and has allowed for the rapid and efficient evaluation of 1280 microbial extracts. The identification of several active compounds validates the potential of the platform for the search for new compounds with antiviral capacity.

microbiology↗

bioRxiv2021

Identification of the 216kbp gene cluster and structure elucidation of gargantulides B and C, new complex 52-membered macrolides from Amycolatopsis sp.

Gargantulides B and C, two new and highly complex 52-membered glycosylated macrolactones, were isolated from Amycolatopsis sp. strain CA-230715 during an antibacterial screening campaign. The structures of these giant macrolides were elucidated by 2D NMR spectroscopy and shown to be related to gargantulide A, although containing additional {beta}-glucopyranose and/or -arabinofuranose monosaccharides separately attached to their backbones. Genome sequencing allowed the identification of a strikingly large 216 kbp biosynthetic gene cluster, among the largest type I PKS clusters described so far, and the proposal of a biosynthetic pathway for gargantulides A-C. Additionally, genes putatively responsible for the biosynthesis of the amino sugar {beta}-3,6-deoxy-3-methylamino glucose, reported exclusively in gargantulide macrolides, were also found in the cluster and described in this work. The absolute configurations of gargantulides B and C were assigned based on a combination of NMR and bioinformatics analysis of ketoreductase and enoylreductase domains within the multimodular type I PKS. Furthermore, the absolute stereochemistry of the related macrolide gargantulide A has now been revised and completed. Gargantulides B and C display potent antibacterial activity against a set of drug-resistant Gram-positive bacteria and moderate activity against the clinically relevant Gram-negative pathogen Acinetobacter baumannii.

microbiology↗