In Silico Design of High-Affinity Antibody VRC34.01 for HIV-1 Fusion Peptide Recognition

The broadly neutralizing antibody VRC34.01, isolated from donor N123, recognizes the N-terminal fusion peptide of the HIV-1 envelope glycoprotein gp41 and inhibits viral membrane fusion. A detailed molecular understanding of this interaction is essential for elucidating the basis of peptide recognition and guiding antibody optimization. Here, molecular dynamics (MD) simulations combined with free energy perturbation (FEP) calculations were used to systematically investigate the effects of point mutations on both the fusion peptide and the antibody. Mutational analyses of key peptide residues showed that most substitutions reduce binding affinity, indicating that the native peptide sequence is finely optimized for recognition by VRC34.01. Decomposition of binding free energies further reveals that these effects are primarily driven by changes in van der Waals interactions associated with hydrophobic packing at the interface. In contrast, in silico mutagenesis of antibody interfacial residues identifies a small subset of affinity-enhancing mutations, including A33I, L96I and S93T. Structural analyses indicate that these mutations improve binding by optimizing local hydrophobic packing without altering the overall binding mode. These results provide a quantitative and structural framework for understanding fusion peptide recognition and highlight a general strategy for antibody affinity optimization through subtle interfacial packing refinement.