Open Access

David Lutje Hulsik, Ying-ying Liu, Nika M. Strokappe, Simone Battella, Mohamed El Khattabi, Laura E. McCoy, Charles Sabin, Andreas Hinz, Miriam Hock, Pauline Macheboeuf, Alexandre M. J. J. Bonvin, Johannes P. M. Langedijk, David Davis, Anna Forsman Quigley, Marlén M. I. Aasa-Chapman, Michael S. Seaman, Alejandra Ramos, Pascal Poignard, Adrien Favier, Jean-Pierre Simorre, Robin A. Weiss, C. Theo Verrips, Winfried Weissenhorn, Lucy Rutten

• The membrane proximal external region (MPER) of the HIV-1 glycoprotein gp41 is targeted by the broadly neutralizing antibodies 2F5 and 4E10. To date, no immunization regimen in animals or humans has produced HIV-1 neutralizing MPER-specific antibodies. We immunized llamas with gp41-MPER proteoliposomes and selected a MPER-specific single chain antibody (VHH), 2H10, whose epitope overlaps with that of mAb 2F5. Bi-2H10, a bivalent form of 2H10, which displayed an approximately 20-fold increased affinity compared to the monovalent 2H10, neutralized various sensitive and resistant HIV-1 strains, as well as SHIV strains in TZM-bl cells. X-ray and NMR analyses combined with mutagenesis and modeling revealed that 2H10 recognizes its gp41 epitope in a helical conformation. Notably, tryptophan 100 at the tip of the long CDR3 is not required for gp41 interaction but essential for neutralization. Thus bi-2H10 is an anti-MPER antibody generated by immunization that requires hydrophobic CDR3 determinants in addition to epitope recognition for neutralization similar to the mode of neutralization employed by mAbs 2F5 and 4E10.