A simple, high-throughput stabilization assay to test HIV-1 uncoating inhibitors
| Autor: | Michaela Rumlová, Kryštof Škach, Alžběta Dostálková, Martin Flegel, Ivana Křížová, Tomáš Ruml, Richard Hrabal, Filip Kaufman, Romana Hadravová |
|---|---|
| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
Anti-HIV Agents 030106 microbiology lcsh:Medicine HIV Infections Article 03 medical and health sciences chemistry.chemical_compound Virus Uncoating Complementary DNA Humans Amino Acid Sequence lcsh:Science Nucleocapsid Retrovirus Multidisciplinary Base Sequence Chemistry Viral Core Proteins lcsh:R High-throughput screening RNA Small molecule Reverse transcriptase High-Throughput Screening Assays Cytosol 030104 developmental biology Capsid HIV-1 Biophysics Nucleic acid RNA Viral lcsh:Q DNA |
| Zdroj: | Scientific Reports Scientific Reports, Vol 9, Iss 1, Pp 1-12 (2019) |
| ISSN: | 2045-2322 |
| Popis: | Shortly after entering the cell, HIV-1 copies its genomic RNA into double-stranded DNA in a process known as reverse transcription. This process starts inside a core consisting of an enclosed lattice of capsid proteins that protect the viral RNA from cytosolic sensors and degradation pathways. To accomplish reverse transcription and integrate cDNA into the host cell genome, the capsid shell needs to be disassembled, or uncoated. Premature or delayed uncoating attenuates reverse transcription and blocks HIV-1 infectivity. Small molecules that bind to the capsid lattice of the HIV-1 core and either destabilize or stabilize its structure could thus function as effective HIV-1 inhibitors. To screen for such compounds, we modified our recently developed FAITH assay to allow direct assessment of the stability of in vitro preassembled HIV-1 capsid-nucleocapsid (CANC) tubular particles. This new assay is a high-throughput fluorescence method based on measuring the amount of nucleic acid released from CANC complexes under disassembly conditions. The amount of disassembled CANC particles and released nucleic acid is proportional to the fluorescence signal, from which the relative percentage of CANC stability can be calculated. We consider our assay a potentially powerful tool for in vitro screening for compounds that alter HIV disassembly. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|