Cryo-EM structures of antibodies elicited by germline-targeting HIV MPER epitope scaffolds

Jiachen Huang; Olivia M Swanson; Kimmo Rantalainen; Monica L Fernández-Quintero; Johannes R Loeffler; Ryan Tingle; Erik Georgeson; Nicole Phelps; Gabriel Ozorowski; Torben Schiffner; William R Schief; Andrew B Ward

doi:10.1016/j.celrep.2025.116818

Cryo-EM structures of antibodies elicited by germline-targeting HIV MPER epitope scaffolds

Cell Rep. 2026 Jan 27;45(1):116818. doi: 10.1016/j.celrep.2025.116818. Epub 2026 Jan 12.

Affiliations

¹ Department of Integrative Structural and Computational Biology, the Scripps Research Institute, La Jolla, CA, USA.
² Department of Integrative Structural and Computational Biology, the Scripps Research Institute, La Jolla, CA, USA; Department of Immunology and Microbiology, the Scripps Research Institute, La Jolla, CA, USA; Consortia for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVD), the Scripps Research Institute, La Jolla, CA, USA; IAVI Neutralizing Antibody Center, the Scripps Research Institute, La Jolla, CA, USA.
³ Department of Immunology and Microbiology, the Scripps Research Institute, La Jolla, CA, USA; Consortia for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVD), the Scripps Research Institute, La Jolla, CA, USA; IAVI Neutralizing Antibody Center, the Scripps Research Institute, La Jolla, CA, USA.
⁴ Department of Integrative Structural and Computational Biology, the Scripps Research Institute, La Jolla, CA, USA; Consortia for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVD), the Scripps Research Institute, La Jolla, CA, USA; IAVI Neutralizing Antibody Center, the Scripps Research Institute, La Jolla, CA, USA.
⁵ Department of Immunology and Microbiology, the Scripps Research Institute, La Jolla, CA, USA; Consortia for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVD), the Scripps Research Institute, La Jolla, CA, USA; IAVI Neutralizing Antibody Center, the Scripps Research Institute, La Jolla, CA, USA; Moderna, Inc., Cambridge, MA, USA. Electronic address: schief@scripps.edu.
⁶ Department of Integrative Structural and Computational Biology, the Scripps Research Institute, La Jolla, CA, USA; Consortia for HIV/AIDS Vaccine Immunology and Immunogen Discovery (CHAVD), the Scripps Research Institute, La Jolla, CA, USA; IAVI Neutralizing Antibody Center, the Scripps Research Institute, La Jolla, CA, USA. Electronic address: andrew@scripps.edu.

PMID: 41528847
DOI: 10.1016/j.celrep.2025.116818

Abstract

Applying cryoelectron microscopy (cryo-EM) to small protein complexes is usually challenging due to their lack of features for particle alignment. Here, we characterized antibody responses to 21 kDa human immunodeficiency virus (HIV) membrane-proximal external region germline-targeting (MPER-GT) immunogens through cryo-EM by complexing them with 10E8 or Fabs derived from MPER-GT-immunized animals. Distinct antibody-antigen interactions were analyzed using atomic models generated from cryo-EM maps. Mutagenesis screening revealed that off-target monoclonal antibodies (mAbs), which do not compete with 10E8, bind non-MPER epitopes, and the binding of two most dominant epitopes were verified by cryo-EM. The structures of 10E8-class on-target Fabs showed binding patterns that resemble the YxFW motif in the 10E8 heavy chain complementarity-determining region 3 (HCDR3) loop. Additionally, we demonstrate that high-resolution maps can be generated from heterogeneous samples with pooled competing Fabs. Overall, our findings will facilitate the optimization of MPER-GT antigens and push the size limit for cryo-EM-based epitope mapping with smaller antigens and heterogeneous antibody mixes.

Keywords: CP: immunology; CP: molecular biology; HIV-1 Env; MPER; cryo-EM; germline targeting; membrane-proximal external region.

MeSH terms

Animals
Antibodies, Monoclonal / immunology
Cryoelectron Microscopy* / methods
Epitopes* / immunology
HIV Antibodies* / chemistry
HIV Antibodies* / immunology
HIV-1* / immunology
Humans
Immunoglobulin Fab Fragments / immunology
Mice
Models, Molecular

Substances

Epitopes
Antibodies, Monoclonal
HIV Antibodies
Immunoglobulin Fab Fragments