Table 1.
Combinatorial alanine scanning results against 4E11A.
| DENV-1 DIIIB |
DENV-2 DIIIB |
||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| Res. | WT/Ala (4E11) |
WT/Ala (M2) |
ΔΔGAla-WT (kcal/mol) |
ΔΔGm2-WT (kcal/mol) |
ΔΔGm3-WT (kcal/mol) |
Res. | WT/Ala (4E11) |
WT/Ala (M2) |
ΔΔGAla-WT (kcal/mol) |
ΔΔGm2-WT (kcal/mol) |
ΔΔGm3-WT (kcal/mol) |
| S305 | 1.0 | 1.4 | −0.18 | - | - | K305 | 5.7 | 2.3 | 0.54 | 0.63 (E) | 0.54 (T) |
| K307 | 4.6 | 3.9 | 0.10 | 0.40 (E) | 0.0 (T) | K307 | 35 | 2.2 | 1.7 | 1.9 (E) | 0.67 (T) |
| L308 | 4.6 | 3.6 | 0.15 | 0.14 (P) | −0.03 (V) | I308 | 15 | 4.7 | 0.70 | 0.65 (T) | 0.45 (V) |
| E309 | 1.5 | 1.4 | 0.04 | - | - | V309 | 1.7 | 2.4 | −0.21 | - | - |
| K310 | 24 | 3.2 | 1.2 | 1.7 (E) | 0.47 (T) | K310 | 11 | 1.9 | 1.0 | 1.6 (E) | 0.90 (T) |
| E311 | 1.5 | 2.8 | −0.36 | - | - | E311 | 2.3 | 2.0 | 0.07 | - | - |
| V312 | 2.7 | 1.6 | 0.29 | - | - | I312 | 20 | 1.8 | 1.4 | 1.5 (T) | 0.46 (V) |
| Q323 | 2.8 | 1.5 | 0.39 | 0.50 (E) | 0.24 (P) | R323 | 0.64 | 0.94 | −0.23 | −0.79 (G) | −0.86 (P) |
| K325 | 3.9 | 2.6 | 0.23 | 1.3 (E) | −1.0 (T) | Q325 | 1.9 | 1.7 | 0.07 | −0.5 (E) | > 2.6 (P)C |
| E327 | 2.3 | 3.0 | −0.16 | - | - | E327 | 6.0 | 1.5 | 0.84 | - | - |
| P332 | 40 | 5.1 | 1.2 | - | - | P332 | 52 | 0.90 | 2.4 | - | - |
| E360 | 2.1 | 1.5 | 0.22 | - | - | ||||||
| K361 | 2.2 | 5.5 | −0.55 | −0.08 (E) | 0.03 (T) | K361 | 1.5 | 0.94 | 0.27 | 0.91 (E) | 0.39 (T) |
| E362 | 1.5 | 1.8 | −0.12 | - | - | D362 | 1.9 | 1.7 | 0.05 | - | - |
| P364 | 0.77 | 0.99 | −0.15 | - | - | P364 | 1.1 | 0.75 | 0.24 | - | - |
| N366 | 0.95 | 1.9 | −0.41 | −0.42 (D) | 0.07 (T) | N366 | 2.3 | 2.5 | −0.05 | 0.25 (D) | 0.24 (T) |
| E384 | 2.4 | 2.3 | 0.01 | - | - | P384 | 7.0 | 0.60 | 1.5 | - | - |
| K385 | 2.9 | 2.2 | 0.16 | 0.36 (E) | 0.13 (T) | G385 | 1.4 | 2.4 | −0.31 | - | - |
| L387 | 18 | 2.6 | 1.1 | 1.2 (P) | 0.15 (V) | L387 | 12 | 0.88 | 1.6 | > 2.7 (P)C | 0.50 (V) |
| K388 | 2.6 | 1.3 | 0.41 | 0.80 (E) | 0.36 (T) | K388 | 0.95 | 1.0 | −0.03 | −0.53 (E) | 0.02 (T) |
| L389 | 8.9 | 2.0 | 0.89 | 2.0 (P) | 0.23 (V) | L389 | 14 | 1.0 | 1.6 | > 2.5 (P)C | 0.39 (V) |
| S390 | 1.5 | 1.1 | 0.18 | - | - | N390 | 2.2 | 1.5 | 0.25 | 1.4 (D) | 0.14 (T) |
| W391 | 5.2 | 1.6 | 0.69 | 0.67 (G) | 0.34 (S) | W391 | 77 | 1.2 | 2.5 | 3.4 (G) | 2.4 (S) |
|
| |||||||||||
| DENV-3 DIIIB |
DENV-4 DIIIB |
||||||||||
| WT/Ala (4E11) |
WT/Ala (M2) |
ΔΔGAla-WT (kcal/mol) |
ΔΔGm2-WT (kcal/mol) |
ΔΔGm3-WT (kcal/mol) |
WT/Ala (4E11) |
WT/Ala (M2) |
ΔΔGAla-WT (kcal/mol) |
ΔΔGm2-WT (kcal/mol) |
ΔΔGm3-WT (kcal/mol) |
||
|
| |||||||||||
| T305 | 0.83 | 0.77 | 0.04 | - | - | K305 | 4.7 | 2.1 | 0.49 | 0.92 (E) | 0.09 (T) |
| V307 | 4.2 | 2.7 | 0.25 | - | - | S307 | 78 | 6.9 | 1.5 | - | - |
| L308 | 32 | 5.9 | 1.0 | > 1.6 (P)C | 1.3 (V) | I308 | >121 | 18 | > 1.2C | > 1.7 (T)C | 0.39 (V) |
| K309 | 0.39 | 1.3 | −0.70 | −0.22 (E) | −0.35 (T) | D309 | 0.01 | 0.58 | −2.3 | - | - |
| K310 | 6.4 | 2.8 | 0.51 | 1.2 (E) | 0.21 (T) | K310 | 157 | 4.8 | 2.1 | > 3.5 (E)C | > 28 (T)C |
| E311 | 3.0 | 1.6 | 0.38 | - | - | E311 | 5.1 | 3.0 | 0.31 | - | - |
| V312 | 16 | 5.3 | 0.67 | - | - | M312 | 10 | 5.0 | 0.43 | > 1.7 (T)C | −0.17 (V) |
| K323 | 8.5 | 3.1 | 0.60 | −0.50 (E) | −0.64 (T) | K323 | NDD | ||||
| E325 | 22 | 7.8 | 0.62 | - | - | K325 | NDD | ||||
| K327 | 52 | 2.5 | 1.8 | > 2.8 (E)C | 1.2 (T) | E327 | NDD | ||||
| P332 | >114 | 1.1 | > 2.8C | - | - | P332 | NDD | ||||
| K361 | 1.4 | 1.2 | 0.08 | 0.59 (E) | −0.25 (T) | T361 | 3.6 | 2.3 | 0.28 | - | - |
| E362 | 2.0 | 2.4 | −0.11 | - | - | N362 | 3.6 | 3.4 | 0.03 | −0.29 (D) | 0.10 (T) |
| P364 | 0.95 | 0.68 | 0.20 | - | - | V364 | 6.5 | 4.4 | 0.23 | - | - |
| N366 | 1.1 | 1.7 | −0.27 | −0.16 (D) | −0.35 (T) | N366 | 1.1 | 0.60 | 0.37 | −0.42 (D) | −0.19 (T) |
| D384 | 1.2 | 1.7 | −0.21 | - | - | D384 | NDD | ||||
| K385 | 5.8 | 3.1 | 0.38 | 0.70 (E) | 0.89 (T) | S385 | NDD | ||||
| L387 | >110 | 1.5 | > 2.6C | > 2.4 (P)C | 2.0 (V) | L387 | NDD | ||||
| K388 | 33 | 1.1 | 2.0 | > 2.8 (E)C | 1.3 (T) | T388 | NDD | ||||
| I389 | > 93 | 1.8 | > 2.4C | > 2.1 (T)C | 0.77 (V) | L389 | NDD | ||||
| N390 | 1.7 | 1.2 | 0.22 | 1.3 (D) | 1.6 (T) | H390 | NDD | ||||
| W391 | 16 | 1.2 | 1.6 | 2.0 (G) | 1.5 (S) | W391 | NDD | ||||
A
Calculated WT/Ala ratios for 4E11 and M2 and ΔΔGAla-WT are listed. In cases where additional residues were permitted, the ΔΔGm2-WT and ΔΔGm3-WT values are also provided. Mutations that were largely deleterious (≥ 1 kcal/mol) are shown in bold.
B
Sample sizes were greater than 100 in most cases (see Supporting Information).
C
In some cases, a single sequence containing a mutation was not isolated in the selected pool, therefore only a lower estimate could be provided.
D
Not determined. Values for positions located in the “lower” segment of the DENV-4 DIII epitope could not be obtained due to inefficient library screening.