Table 1 Mouse TfR binding nanobody histidine mutants tested in vitro and in vivo
Nanobody | mTfR affinity pH 7.4 (nM) | Apparent mTfR unbinding ratio at pH 5.5 vs pH 7.4 | NT-mediated Hypothermia (°C) |
|---|---|---|---|
M1 | 5.11 | 1.01 | 0 |
M1: P96H | 7.031 | 1.46 | 1 |
M1: Y98H | 2509 | 1.28 | 2.5 |
M1: S99H | > 5000 | 1.01 | 1 |
M1: T100aH | 55.31 | 1.05 | 1.5 |
M1: T100bH | 30.12 | 1.24 | 0 |
M1: R100dH | 1294 | 1.01 | 0 |
M1: L100eH | 203 | 1.32 | 0 |
M1: D100fH | > 5000 | 1.58 | 0 |
M1: S100gH | > 5000 | 1.03 | 0 |
M1: K100iH | > 5000 | 1.42 | 0 |
M1: D101H | 25 | 1.32 | 1.5 |
M1: Y32H, P96H | 373 | 2.57 | 0 |
M1: G65H, P96H | > 5000 | 2.09 | 1 |
M1: P96H, Y98H | 33.18 | 2.48 | 1.2 |
M1: P96H, S100H | 1140 | 1.02 | 0 |
M1: P96H, D101H | 242 | 2.65 | 7.5 |
M1: P96H, Y102H | 11.65 | 2.33 | 6 |
M1: Y98H, Y102H | > 5000 | 1.29 | 5 |
M1: D31H P96H Y102H | 95 | 1.13 | 0 |
M1: R33H P96H Y102H | 1545 | 1.10 | 0 |
M1: R56H, P96H, Y98H | > 5000 | 0.98 | 1.5 |
M1: R56H, P96H, D101H | > 5000 | 1.78 | 0 |
M1: R56H, P96H, Y102H | 6.653 | 2.63 | 8 |
M1: R56H, Y98H, D101H | 52.6 | 1.04 | 0 |
M1: R56H, Y98H, Y102H | > 5000 | 0.99 | 0 |
M1: R56A, P96H, Y102H | 1.7 | 2.59 | 0 |
M1: Y59H, P96H, Y102H | 888 | 2.07 | 1 |
M1: A60H, P96H, Y102H | > 5000 | ND | 0 |
M1: P61H, P96H, Y102H | 148 | 2.13 | 1 |
M1: S62H, P96H, Y102H | > 5000 | 2.18 | 1.5 |
M1: A64H, P96H, Y102H | > 5000 | 1.79 | 0 |
M1: S70H, P96H, Y102H | 78 | 1.64 | 0 |
M1: P96H S99H D101H | 301 | 2.43 | 2 |
M1: P96H S99H Y102H | > 5000 | 2.52 | 2.5 |
M1: P96H, D101H, Y102H | > 5000 | 2.42 | 0 |
M1: C22A C92A P96H D101H | 35.76 | 1.03 | 0 |
M1: C22A C92A P96H Y102H | 17.56 | 1.07 | 0 |
M1: R56H, P96H, Y98H, Y102H | > 5000 | 1.00 | 0 |
M1: R56H, P96H, S99H, Y102H | 246 | 2.04 | 5.5 |
M1: Y32H, R56H, P96H, S99H, Y102H |  > 5000 | 1.11 | 0 |