Table 3.
Distribution of hotspot paratope residues from prior alanine-scanning antibody studies.
| Ab/Ag | Method of hotspot determination | Number of paratope positions analyzed | Crystal structure | HCDR1 | HCDR2 | HCDR3 | VH | LCDR1 | LCDR2 | LCDR3 | VL |
|---|---|---|---|---|---|---|---|---|---|---|---|
| m102.4/NiV GP | Ala-scan + ELISA | 28 | N/A | 0 | 0 | 2 | 2 | 0 | 0 | 0 | 0 |
| Y0317/VEGF | Phage display + competitive ELISA | 68 | 1CZ8 | 5 | 3 | 6 | 14 | 0 | 0 | 2 | 2 |
| HyHEL-10/HEL | Ala-scan + ELISA | 12 | 2DQJ | 1 | 3 | 1 | 5 | 2 | 1 | 1 | 4 |
| b12/HIV gp120 | Ala-scan + ELISA | 18 | 1HZH | N/A | N/A | 7 | 7 | N/A | N/A | N/A | 0 |
| 2F5/HIV gp41 | Ala-scan + ELISA | 22 | 2F5B | N/A | N/A | 4 | 4 | N/A | N/A | N/A | 0 |
| Fab2C4/ErbB2 | Phage display + ELISA | 61 | 1L7I | 4 | 8 | 7 | 19 | 0 | 1 | 1 | 2 |
| 6B4/GP lba | Ala-scan + ELISA | 12 | N/A | N/A | 0 | 1 | 1 | 4 | N/A | 1 | 5 |
| N2/alpha-synuclein | Ala-scan + ELISA | 18 | 2X6M | N/A | 4 | 2 | 6 | N/A | N/A | N/A | N/A |
| Fab 13B8.2/CD4 | Spot peptide Ala-scan | 16 | TBD | 1 | 1 | 0 | 2 | 0 | 0 | 0 | 0 |
| HA22-LR/CD22 | Ala-scan + WST-8 assay | 36 | N/A | 0 | N/A | 4 | 4 | 1 | 0 | 0 | 1 |
| HzKR127/HepB | Ala-scan + ELISA | 50 | 2EH8 | 3 | 3 | 2 | 8 | 7 | 1 | 6 | 14 |
| 82D6A3/VWF | Ala-scan + ELISA | 13 | 2ADF | 0 | 0 | 2 | 2 | 0 | 0 | 0 | 0 |
| Anti-PAI | Ala-scan + SPR (not only single mutants) | 26 | N/A | 1 | 0 | 3 | 4 | 0 | 0 | 5 | 5 |
| Fab37/HER2 | Phage display, saturation mutagenesis | 20 | 3N85 | 3 | 1 | 4 | 8 | N/A | N/A | 4 | 4 |
| bH1/VEGF | Ala-scan + SPR | 35 | 3BE1 | 0 | 0 | 2 | 2 | 2 | 0 | 1 | 3 |
| Cetuximab/EGFR | Ala-scan + SPR | 27 | 1YY9 | 0 | 1 | 2 | 3 | 0 | 0 | 1 | 1 |
| D1.3/HEL | Ala-scan + SPR | 10 | 1DVF | 0 | 0 | 2 | 2 | 0 | 0 | 1 | 1 |
Criteria for hotspot determination were for a nearly tenfold impact of mutation on Kd (equivalent to a 1.36 kcal/mol ddG difference). “N/A” refers to CDR loops that were not scanned in the analysis.