Mechanisms of recognition of amyloid-β (Aβ) monomer, oligomer, and fibril by homologous antibodies

Abstract

Alzheimer's disease is one of the most devastating neurodegenerative diseases without effective therapies. Immunotherapy is a promising approach, but amyloid antibody structural information is limited. Here we simulate the recognition of monomeric, oligomeric, and fibril amyloid-β (Aβ) by three homologous antibodies (solanezumab, crenezumab, and their chimera, CreneFab). Solanezumab only binds the monomer, whereas crenezumab and CreneFab can recognize different oligomerization states; however, the structural basis for this observation is not understood. We successfully identified stable complexes of crenezumab with Aβ pentamer (oligomer model) and 16-mer (fibril model). It is noteworthy that solanezumab targets Aβ residues 16-26 preferentially in the monomeric state; conversely, crenezumab consistently targets residues 13-16 in different oligomeric states. Unlike the buried monomeric peptide in solanezumab's complementarity-determining region, crenezumab binds the oligomer's lateral and edge residues. Surprisingly, crenezumab's complementarity-determining region loops can effectively bind the Aβ fibril lateral surface around the same 13-16 region. The constant domain influences antigen recognition through entropy redistribution. Different constant domain residues in solanezumab/crenezumab/chimera influence the binding of Aβ aggregates. Collectively, we provide molecular insight into the recognition mechanisms facilitating antibody design.

Keywords: Alzheimer disease; amyloid; amyloid-beta (AB); antibody; immunotherapy; protein dynamic.

Figure 1.

The structure and dynamics of the three Fabs in their apo form suggest that when unbound to antigens, solanezumab is more conformationally diverse than the other two Fabs. a, 2D RMSDs of the structures sampled in the MD simulation. The structures of crenezumab and CreneFab are taken from PDB entries 4KMV (A) and 4KNA (B), respectively. b, cluster analysis, with backbone RMSD = 4 Å, used to define the cluster. Clusters are colored blue, red, and gray, respectively. c, motion correlation among the residues of the three Fabs. Residues with highly correlated and anti-correlated motion are red and blue, respectively. d, RMSFs of the three Fabs from five independent MD simulations. The RMSFs of Smab0, CHmab0a, CHmab0b, Cmab0a, and Cmab0b are colored black, red, pink, blue, and cyan, respectively.

Figure 2.

The V domains of crenezumab and CreneFab can recognize a more diverse Aβ ensemble than solanezumab. a, the two Aβ-bound crystal structures superimposed on the Aβ and V domains. 4XXD and 4KNA are colored red and lime, respectively. b, clustered conformations of Aβ12–28 in complex with solanezumab, crenezumab, and CreneFab (RSMD for clustering is 4 Å). c, secondary structure components of Aβ12–28. Helical, β-strand, turn, and random coil structures are colored black, red, blue, and green, respectively. d, contact preference on the Fabs–Aβ interface from the Aβ and Fab sides. The Fab amino acid preference was obtained by summation of the contacts based on the Fab side contact preference.

Figure 3.

Solanezumab is less conformationally diverse than the other two Fabs when bound to Aβ12–28. a, cluster analysis of the structures. Backbone RMSD = 4 Å is used to define the cluster. Clusters are colored blue, red, and gray, respectively. b, motion correlation among the residues of the three Fabs. Residues with highly correlated and anti-correlated motion are red and blue, respectively. c and d, RMSFs (c) and order parameters S² (d) of the three Fabs in complex with Aβ12–28. The location of CDRs and important constant domain loops are boxed. The values of solanezumab, crenezumab, and CreneFab are colored black, blue, and red, respectively.

Figure 4.

Flexibility and structural analysis of Aβ monomer, oligomers, and fibrils. a, RMSFs of the three Aβ aggregates. Shown are structures or clustered structures (b) and secondary structure components (c) of Aβ monomer, 5-mer, and 16-mer, respectively. Helical, β, turn, and random coil structures are colored black, red, blue, and green, respectively.

Figure 5.

The epitope of Aβ oligomer shifted to the N-terminal hydrophilic and cationic residues when in complex with crenezumab and CreneFab compared with solanezumab. a, molecular details of the Aβ oligomer-crenezumab complex. Residues with cumulative contacts >1.0 are represented by sticks. Crenezumab residues Tyr²⁵¹H and Asp³¹⁹H (alanine scanning using SPR (23)) are highlighted by beads, whereas other important residues, Ser²⁵⁰H and Asn³³L, which differ between crenezumab and solanezumab, are also underlined. Residues from Aβ, light chain, and heavy chain are indicated by Aβ, L, and H, respectively. Light chain, heavy chain, and Aβ oligomer are colored pink, lime, and ice blue, respectively. Hydrophobic, hydrophilic, cationic, and anionic residues are colored white, green, blue, and red, respectively. b, contact preference on the Fabs–Aβ interface from Aβ side and Fab side. Fab amino acids preference was obtained by summation of the contacts based on the Fab side contact preference.

Figure 6.

Partially ordered Aβ oligomers induce the subdomain reorientation of the crenezumab Fab to transfer the entropy upon stable antibody–antigen interface formation. a, cluster analysis of the structures. Backbone RMSD = 4 Å is used to define the cluster. Clusters are colored blue, red, gray, yellow, and orange, respectively. b, motion correlation among the residues of the three Fabs and Aβ oligomer. Residues with highly correlated or anti-correlated motion are red or blue. c and d, RMSFs (c) and order parameters S² (d) of the three Fabs in complex with Aβ oligomer. The locations of CDRs and important constant domain loops are boxed. The curves of solanezumab, crenezumab, and CreneFab are colored black, blue, and red, respectively.

Figure 7.

The array of N-terminal hydrophilic and cationic residues of Aβ fibrils were recognized by crenezumab with dominant salt bridges and hydrogen bonds. a, molecular details of the Aβ fibrils-crenezumab complex. Residues with cumulative contacts >1.0 are represented by sticks. Crenezumab residues Tyr²⁵¹H (alanine scanning using SPR (23)) are further highlighted by beads, whereas Asn²⁷³H, Asp³⁵L, and Asn³³L, which differ between crenezumab and solanezumab, are also underlined. Residues from Aβ, light chain, and heavy chain are indicated by Aβ, L, and H, respectively. Light chain, heavy chain, and Aβ oligomer are colored pink, lime, and ice blue, respectively. Hydrophobic, hydrophilic, cationic, and anionic residues are colored white, green, blue, and red, respectively. b, contact preference on the Fabs–Aβ interface from the Aβ side and Fab side. Fab amino acid preference was obtained by summation of the contacts based on the Fab side contact preference.

Figure 7.

The array of N-terminal hydrophilic and cationic residues of Aβ fibrils were recognized by crenezumab with dominant salt bridges and hydrogen bonds. a, molecular details of the Aβ fibrils-crenezumab complex. Residues with cumulative contacts >1.0 are represented by sticks. Crenezumab residues Tyr²⁵¹H (alanine scanning using SPR (23)) are further highlighted by beads, whereas Asn²⁷³H, Asp³⁵L, and Asn³³L, which differ between crenezumab and solanezumab, are also underlined. Residues from Aβ, light chain, and heavy chain are indicated by Aβ, L, and H, respectively. Light chain, heavy chain, and Aβ oligomer are colored pink, lime, and ice blue, respectively. Hydrophobic, hydrophilic, cationic, and anionic residues are colored white, green, blue, and red, respectively. b, contact preference on the Fabs–Aβ interface from the Aβ side and Fab side. Fab amino acid preference was obtained by summation of the contacts based on the Fab side contact preference.

Figure 8.

Recognition of highly ordered Aβ fibrils required more flexible and dynamic Fabs to transfer the entropy from antibody–antigen complex formation. a, cluster analysis of the structures. Backbone RMSD = 4 Å is used to define the cluster. Clusters are colored blue, red, and gray, respectively. b, motion correlation among the residues of the three Fabs and Aβ fibril. Residues with highly correlated and anti-correlated motion are red and blue. c and d, RMSFs (c) and order parameters S² (d) of the three Fabs in complex with Aβ fibril. The locations of CDRs and important constant domain loops are boxed. The curves of solanezumab, crenezumab, and CreneFab are colored black, blue, and red, respectively.

Figure 9.

Analysis of RMSFs (a) and order parameters S² of Fab residues (b) in the different simulation systems suggested the transfer of entropy from CDRs to constant domain loops. The RMSFs/order parameters of Fabs in apo form and in complex with Aβ monomer, pentamer, and 16-mer are colored black, red, green, and blue, respectively.