title: Conformational and structural features of HIV-1 gp120 underlying the dual receptor antagonism by cross-reactive neutralizing antibody m18.
abstract: We investigated the interaction between cross-reactive HIV-1 neutralizing human monoclonal antibody m18 and HIV-1YU-2 gp120 in an effort to understand how this antibody inhibits the entry of virus into cells. m18 binds to gp120 with high affinity (KD≈5 nM) as measured by surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC). SPR analysis further showed that m18 inhibits interactions of gp120 with both soluble CD4 and CD4-induced antibodies that have epitopes overlapping the coreceptor binding site. This dual receptor site antagonism, which occurs with equal potency for both inhibition effects, argues that m18 is not functioning as a mimic of CD4, in spite of the presence of a putative CD4-like loop formed by HCDR3 in the antibody. Consistent with this view, m18 was found to interact with gp120 in the presence of saturating concentrations of a CD4-mimicking small molecule gp120 inhibitor, suggesting that m18 does not require unoccupied CD4 Phe43 binding cavity residues of gp120. Thermodynamic analysis of the m18-gp120 interaction suggests that m18 stabilizes a conformation of gp120 that is unique from and less structured than the CD4-stabilized conformation. Conformational mutants of gp120 were studied for their impact on m18 interaction. Mutations known to disrupt the coreceptor binding region and to lead to complete suppression of 17b binding had minimal effects on m18 binding. This argues that energetically important epitopes for m18 binding lie outside the disrupted bridging sheet region used for 17b and coreceptor binding. In contrast, mutations in the CD4 region strongly affected m18 binding. Overall, the results obtained in this work argue that m18, rather than mimicking CD4 directly, suppresses both receptor binding site functions of HIV-1 gp120 by stabilizing a nonproductive conformation of the envelope protein. These results can be related to prior findings about the importance of conformational entrapment as a common mode of action for neutralizing CD4bs antibodies, with differences mainly in epitope utilization and the extent of gp120 structuring.
authors.affiliation: Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania 19102, United States.
authors.forename: Syna Kuriakose
authors.initials: SK
authors.lastname: Gift
authors.forename: Isaac J
authors.initials: IJ
authors.lastname: Zentner
authors.forename: Arne
authors.initials: A
authors.lastname: Schön
authors.forename: Karyn
authors.initials: K
authors.lastname: McFadden
authors.forename: M
authors.initials: M
authors.lastname: Umashankara
authors.forename: Srivats
authors.initials: S
authors.lastname: Rajagopal
authors.forename: Mark
authors.initials: M
authors.lastname: Contarino
authors.forename: Caitlin
authors.initials: C
authors.lastname: Duffy
authors.forename: Joel R
authors.initials: JR
authors.lastname: Courter
authors.forename: Mei-Yun
authors.initials: MY
authors.lastname: Zhang
authors.forename: Jonathan M
authors.initials: JM
authors.lastname: Gershoni
authors.forename: Simon
authors.initials: S
authors.lastname: Cocklin
authors.forename: Dimiter S
authors.initials: DS
authors.lastname: Dimitrov
authors.forename: Amos B
authors.initials: AB
authors.lastname: Smith
authors.forename: Ernesto
authors.initials: E
authors.lastname: Freire
authors.forename: Irwin M
authors.initials: IM
authors.lastname: Chaiken
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism; D057134:Antibodies, Neutralizing / Q000276:immunology / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D015704:CD4 Antigens / Q000276:immunology / Q000378:metabolism; D002151:Calorimetry; D000939:Epitopes / Q000276:immunology / Q000378:metabolism; D015483:HIV Antibodies / Q000276:immunology / Q000378:metabolism; D015699:HIV Envelope Protein gp120 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015497:HIV-1 / Q000276:immunology / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D020349:Surface Plasmon Resonance; D013816:Thermodynamics
chemical_list: D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D015704:CD4 Antigens; D000939:Epitopes; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120