Method to identify efficiently cleaved, membrane-bound, functional HIV-1 (Human Immunodeficiency Virus-1) envelopes

Area of Science:

• Virology
• Immunology
• Vaccine Development

Background:

• An ideal HIV-1 vaccine should elicit broadly neutralizing antibodies (bNAbs) capable of neutralizing diverse viral strains.
• Current vaccine strategies focus on HIV-1 envelope (Env) proteins as immunogens to induce bNAbs.
• Efficient cleavage of the HIV-1 Env precursor (gp160) into subunits is critical for desirable antigenic properties.

Purpose of the Study:

• To present a detailed six-step laboratory method for identifying efficiently cleaved HIV-1 Envs.
• To enable the selection of suitable Env proteins from various clades for immunogen design.
• To facilitate the development of HIV-1 vaccines that elicit potent bNAbs.

Main Methods:

• Optimization of existing assays: FACS-based cell surface antibody binding, pseudovirus neutralization, and gp120 shedding assays.
• Development of new protocols: Biotinylation-neutravidin-agarose pull-down and plasma membrane protein immunoprecipitation assays.
• A repeatable six-step process for characterizing membrane-bound, functional HIV-1 Envs.

Main Results:

• A robust six-step protocol for identifying efficiently cleaved HIV-1 Envs was established.
• The method demonstrated high repeatability for characterizing Env proteins.
• New immunoprecipitation techniques were validated for assessing Env cleavage.

Conclusions:

• The described protocols provide a valuable resource for the scientific community to identify suitable HIV-1 Envs for vaccine development.
• This methodology can be applied to characterize any HIV-1 envelope protein.
• The identification of efficiently cleaved Envs is a key step towards designing effective HIV-1 vaccines eliciting bNAbs.