title: A yeast glycoprotein shows high-affinity binding to the broadly neutralizing human immunodeficiency virus antibody 2G12 and inhibits gp120 interactions with 2G12 and DC-SIGN.
abstract: The human immunodeficiency virus type 1 (HIV-1) envelope (Env) protein contains numerous N-linked carbohydrates that shield conserved peptide epitopes and promote trans infection by dendritic cells via binding to cell surface lectins. The potent and broadly neutralizing monoclonal antibody 2G12 binds a cluster of high-mannose-type oligosaccharides on the gp120 subunit of Env, revealing a conserved and highly exposed epitope on the glycan shield. To find an effective antigen for eliciting 2G12-like antibodies, we searched for endogenous yeast proteins that could bind to 2G12 in a panel of Saccharomyces cerevisiae glycosylation knockouts and discovered one protein that bound weakly in a Delta pmr1 strain deficient in hyperglycosylation. 2G12 binding to this protein, identified as Pst1, was enhanced by adding the Delta mnn1 deletion to the Delta pmr1 background, ensuring the exposure of terminal alpha1,2-linked mannose residues on the D1 and D3 arms of high-mannose glycans. However, optimum 2G12 antigenicity was found when Pst1, a heavily N-glycosylated protein, was expressed with homogenous Man(8)GlcNAc(2) structures in Delta och1 Delta mnn1 Delta mnn4 yeast. Surface plasmon resonance analysis of this form of Pst1 showed high affinity for 2G12, which translated into Pst1 efficiently inhibiting gp120 interactions with 2G12 and DC-SIGN and blocking 2G12-mediated neutralization of HIV-1 pseudoviruses. The high affinity of the yeast glycoprotein Pst1 for 2G12 highlights its potential as a novel antigen to induce 2G12-like antibodies.
authors.affiliation: ProSci Inc., 12170 Flint Place, Poway, CA 92064, USA.
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Luallen
authors.forename: Hu
authors.initials: H
authors.lastname: Fu
authors.forename: Caroline
authors.initials: C
authors.lastname: Agrawal-Gamse
authors.forename: Innocent
authors.initials: I
authors.lastname: Mboudjeka
authors.forename: Wei
authors.initials: W
authors.lastname: Huang
authors.forename: Fang-Hua
authors.initials: FH
authors.lastname: Lee
authors.forename: Lai-Xi
authors.initials: LX
authors.lastname: Wang
authors.forename: Robert W
authors.initials: RW
authors.lastname: Doms
authors.forename: Yu
authors.initials: Y
authors.lastname: Geng
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000378:metabolism; D017951:Antigen Presentation; D000080908:Broadly Neutralizing Antibodies; D015815:Cell Adhesion Molecules / Q000276:immunology* / Q000378:metabolism; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D006023:Glycoproteins / Q000276:immunology* / Q000378:metabolism; D015483:HIV Antibodies / Q000276:immunology* / Q000378:metabolism; D015699:HIV Envelope Protein gp120 / Q000276:immunology* / Q000378:metabolism; D015497:HIV-1 / Q000276:immunology / Q000378:metabolism; D006801:Humans; D037181:Lectins, C-Type / Q000276:immunology* / Q000378:metabolism; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000276:immunology* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000276:immunology / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000276:immunology* / Q000378:metabolism; D020349:Surface Plasmon Resonance; D000099215:DC-Specific ICAM-3 Grabbing Nonintegrin
chemical_list: D000911:Antibodies, Monoclonal; D000080908:Broadly Neutralizing Antibodies; D015815:Cell Adhesion Molecules; D006023:Glycoproteins; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; D037181:Lectins, C-Type; D011134:Polysaccharides; D011956:Receptors, Cell Surface; D029701:Saccharomyces cerevisiae Proteins; D000099215:DC-Specific ICAM-3 Grabbing Nonintegrin; C529157:2G12 monoclonal antibody; C499369:PST1 protein, S cerevisiae