Structural and Thermodynamic Basis of Epitope Binding by Neutralizing and Nonneutralizing Forms of the Anti-HIV-1 Antibody 4E10.
Bottom Line: The conclusions of our structure-function analysis strengthen the idea that to exert effective neutralization, the hydrophobic apex of the solvent-exposed CDR-H3 loop must recognize an antigenic structure more complex than just the linear α-helical epitope and likely constrained by the viral membrane lipids.However, 4E10 (or 4E10-like) antibodies are rarely found in HIV-1-infected individuals or elicited through vaccination.We conclude that the difference between neutralizing and nonneutralizing antibodies of 4E10 is neither structural nor energetic but is related to the capacity to recognize the HIV-1 gp41 epitope inserted in biological membranes.
Affiliation: Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, Tokyo, Japan Biophysics Unit (CSIC, UPV/EHU) and Department of Biochemistry and Molecular Biology, University of the Basque Country (UPV/EHU), Bilbao, Spain.Show MeSH
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Mentions: The results in Fig. 8 strengthen the conclusion that availability of the WT CDR-H3 apex contributes to additional interactions with membrane components as previously suggested (10, 14, 22, 45–48). The DNA vaccine candidates, MPER-TM1 and MPER-PDGFR (22), express fusion proteins comprising: (i) an N-terminal HA tag, (ii) the MPER, (iii) either the transmembrane domain of gp41 followed by 27-AA of the cytoplasmic domain (TM1) or that of the platelet-derived growth-factor receptor (PDGFR). These fusions are expressed in the context of the plasma membrane; however, the PDGFR transmembrane domain has been shown to reduce exposure of the 4E10 epitope (22) and, to a lesser extent, the 10E8 epitope (N. Gulzar, unpublished data). The titrations shown in Fig. 8A indicate that IgG and Fab WT engaged effectively with the 4E10 epitope presented in the context of the plasma membrane (EC50s of ca. 0.1 and 1 nM, respectively). In comparison, in the range of measured concentrations, binding to MPER-TM1 was negligible for the WDWD and ΔLoop mutants (Fig. 8A, bottom panels). None of the antibodies bound directly to the plasma membrane of cells devoid of an MPER construct (squares and dotted lines), demonstrating that the binding scored for the WT IgG and Fab under our experimental conditions was strictly dependent on the presence of the MPER epitope.
Affiliation: Department of Bioengineering, Graduate School of Engineering, The University of Tokyo, Tokyo, Japan Biophysics Unit (CSIC, UPV/EHU) and Department of Biochemistry and Molecular Biology, University of the Basque Country (UPV/EHU), Bilbao, Spain.