title: HIV-1 gp120 dimers decrease the overall affinity of gp120 preparations for CD4-induced ligands.
abstract: For several years, tools to study the conformational changes of HIV-1 envelope glycoproteins have been developed in order to comprehend those changes and their role in the fusion process and immunogenicity of HIV-1. To facilitate these studies, expression of the HIV-1 gp120 envelope glycoprotein has been done in several over-expression settings. However, over-expression of HIV-1 gp120 in mammalian cells leads to the formation of aberrant disulfide-linked dimers that can bias the results of experiments aimed at measuring gp120 affinity with different ligands. The presence of these gp120 dimers, generated in a widely used gp120 expression system, affects the affinity of gp120 for CD4-induced ligands, as evaluated by surface plasmon resonance. Upon monomeric gp120 purification, neither the removal of potential glycosylation sites on V4 nor the removal of the V5 variable region affect the overall affinity of gp120 for 17b and A32 CD4-induced ligands. Removal of these aberrant disulfide-linked gp120 dimers by standard size exclusion chromatography is sufficient to restore the overall affinity of gp120 preparations for these ligands.
authors.affiliation: Centre de Recherche du CHUM, Université de Montréal, Montreal, Quebec, Canada; Department of Microbiology, Infectiology and Immunology, Université de Montréal, Montreal, Quebec, Canada.
authors.forename: Mathieu
authors.initials: M
authors.lastname: Coutu
authors.affiliation: Centre de Recherche du CHUM, Université de Montréal, Montreal, Quebec, Canada; Department of Microbiology, Infectiology and Immunology, Université de Montréal, Montreal, Quebec, Canada; Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada. Electronic address: andres.finzi@umontreal.ca.
authors.forename: Andrés
authors.initials: A
authors.lastname: Finzi
mesh_terms: D015704:CD4 Antigens / Q000378:metabolism*; D002850:Chromatography, Gel / Q000379:methods; D015699:HIV Envelope Protein gp120 / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D011485:Protein Binding; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D020349:Surface Plasmon Resonance
chemical_list: D015704:CD4 Antigens; D015699:HIV Envelope Protein gp120; D011994:Recombinant Proteins; C514793:gp120 protein, Human immunodeficiency virus 1
keywords: CD4-induced epitopes; FPLC; HIV; Surface plasmon resonance; gp120; gp120 dimers