The Highly Conserved Glycan at Asparagine 260 of HIV-1 gp120 Is Indispensable for Viral Entry

Abstract

Carbohydrate-binding agents bind to the N-glycans of HIV-1 envelope gp120 and prevent viral entry. Carbohydrate-binding agents can select for mutant viruses with deleted envelope glycans. Not all glycosylation motifs are mutated to the same extent. Site-directed mutagenesis revealed that deletions destroying the highly conserved 260 NGS 262 glycosylation motif resulted in non-infectious virus particles. We observed a significant lower CD4 binding in the case of the N260Q mutant gp120 virus strains, caused by a strikingly lower expression of gp120 and gp41 in the virus particle. In addition, the mutant N260Q HIV-1 envelope expressed in 293T cells was unable to form syncytia in co-cultures with U87.CD4.CXCR4.CCR5 cells, due to the lower expression of envelope protein on the surface of the transfected 293T cells. The detrimental consequence of this N-glycan deletion on virus infectivity could not be compensated for by the creation of novel glycosylation sites near this amino acid, leaving this uncovered envelope epitope susceptible to neutralizing antibody binding. Thus, the Asn-260 glycan in the gp120 envelope of HIV-1 represents a hot spot for targeting suicidal drugs or antibodies in a therapeutic effort to efficiently neutralize a broad array of virus strains.

Mutation of a Single Envelope N-Linked Glycosylation Site Enhances the Pathogenicity of Bovine Leukemia Virus

“…Use of pharmacological inhibitors and lectins suggested that, as expected, glycosylation is involved in cell fusion, confirming and expanding previous reports (26,31,32) (Fig. 1).…”

HIV-1 Envelope Glycan Moieties Modulate HIV-1 Transmission

“…N-Glycans include three basic structures, namely, high-mannose (i.e., oligomannose), hybrid, and complex glycans, all of which are present on HIV-1 Env (4-7). Importantly, N-glycans shield the Env backbone and have been shown to play key roles in determining the Env structure, epitope exposure, and, consequently, antigenicity, immunogenicity, antibody neutralization, infectivity, and carbohydrate and receptor binding (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20).…”

“…N-Glycans include three basic structures, namely, high-mannose (i.e., oligomannose), hybrid, and complex glycans, all of which are present on HIV-1 Env (4-7). Importantly, N-glycans shield the Env backbone and have been shown to play key roles in determining the Env structure, epitope exposure, and, consequently, antigenicity, immunogenicity, antibody neutralization, infectivity, and carbohydrate and receptor binding (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20).The position of N-linked glycans on glycoproteins is genetically encoded, whereas the types of glycans at a given Asn in the consensus Asn-X-Ser/Thr-X sequence (where X is not Pro) are determined by the glycan branching processes in the cell endoplasmic reticulum and Golgi apparatus (21). However, some glycoproteins, including HIV-1 gp120, demonstrate protein-specific glycosylation (22)(23)(24)(25).…”

“…Studies of HIV-1 glycosylation have focused mainly on the Env structure and surface exposure and their effect on antigenicity, immunogenicity, and antibody neutralization (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20) through mutation of the position of glycosylation, rather than modification of the type of glycans. Studies of the effect of glycosylation on HIV-1 infectivity and carbohydrate and receptor binding have also utilized deletion of the potential N-linked glycosylation sites (PNGSs) (15)(16)(17)(18)28).…”

Entry Inhibitors Directed Towards Glycoprotein gp120: An Overview on a Promising Target for HIV-1 Therapy

“…Moreover, it has been reported that glycan deletions in the HIV-1 gp120 V1/V2 domains severely compromise viral infectivity and sensitize the mutant virus strains to carbohydrate-binding agents [56]. Furthermore, some specific Nglycosylation sites are indispensable for efficient viral entry (such as Asn 260 ) [57]. Thus, a resistance pattern directed to the elimination of N-glycosylation sites of gp120 could be of interest in enhancing immunosuppression of the pathogen.…”