A Fluorescence Polarization Assay for High-Throughput Screening of Inhibitors against HIV-1 Nef-Mediated MHC-I Downregulation.
J Biol Chem
; : 107529, 2024 Jul 01.
Article
en En
| MEDLINE
| ID: mdl-38960039
ABSTRACT
The multifunctional, HIV-1 accessory protein Nef enables infected cells to evade host immunity and thus plays a key role in viral pathogenesis. One prominent function of Nef is the downregulation of major histocompatibility complex class I (MHC-I), which disrupts antigen presentation and thereby allows the infected cells to evade immune surveillance by the cytotoxic T cells. Therapeutic inhibition of this Nef function is a promising direction of antiretroviral drug discovery as it may revitalize cytotoxic T cells to identify, and potentially clear, hidden HIV-1 infections. Guided by the crystal structure of the protein complex formed between Nef, MHC-I, and the hijacked clathrin adaptor protein complex 1 (AP1), we have developed a fluorescence polarization-based assay for inhibitor screening against Nef's activity on MHC-I. The optimized assay has a good signal-to-noise ratio, substantial tolerance of DMSO, and excellent ability to detect competitive inhibition, indicating that it is suitable for high-throughput screening.
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01-internacional
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MEDLINE
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En
Revista:
J Biol Chem
Año:
2024
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Article