ITC-binding measurements of the HKD5 interaction with gC1qR
| . | Binding model . | N . | KD (nM) . | ΔH (kJ mol−1) . | ΔS (J K−1 M−1) . | ΔG (kJ mol−1) . |
|---|---|---|---|---|---|---|
| D5 | 3-site sequential binding | — | 1) 1.9 (±0.1) | 1) −268.8 (±1.1) | 1) −732.7 | 1) −50.5 |
| 2) 64.9 (±1.9) | 2) −105.2 (±1.3) | 2) −214.8 | 2) −41.2 | |||
| 3) 1011.1 (±75.8) | 3) −23.5 (±1.4) | 3) 36.0 | 3) −34.3 | |||
| D5-1 | 3-site sequential binding | — | 1) 73.0 (±4.0) | 1) −200.9 (±1.4) | 1) −535.9 | 1) −41.2 |
| 2) 1579.8 (±145.3) | 2) −92.2 (±4.9) | 2) −198.5 | 2) −33.0 | |||
| 3) 2673.8 (±257.7) | 3) −62.2 (±5.9) | 3) −101.7 | 3) −31.9 | |||
| D5-2 | Single site | 2.30 (±0.01) | 763.4 (±27.5) | −1459.1 (±1.1) | −382.7 | −35.1 |
| HK 496-516 | Single site | 2.56 (±0.02) | 1612.9 (±90.3) | −164.0 (±1.8) | −439.6 | −33.2 |
| . | Binding model . | N . | KD (nM) . | ΔH (kJ mol−1) . | ΔS (J K−1 M−1) . | ΔG (kJ mol−1) . |
|---|---|---|---|---|---|---|
| D5 | 3-site sequential binding | — | 1) 1.9 (±0.1) | 1) −268.8 (±1.1) | 1) −732.7 | 1) −50.5 |
| 2) 64.9 (±1.9) | 2) −105.2 (±1.3) | 2) −214.8 | 2) −41.2 | |||
| 3) 1011.1 (±75.8) | 3) −23.5 (±1.4) | 3) 36.0 | 3) −34.3 | |||
| D5-1 | 3-site sequential binding | — | 1) 73.0 (±4.0) | 1) −200.9 (±1.4) | 1) −535.9 | 1) −41.2 |
| 2) 1579.8 (±145.3) | 2) −92.2 (±4.9) | 2) −198.5 | 2) −33.0 | |||
| 3) 2673.8 (±257.7) | 3) −62.2 (±5.9) | 3) −101.7 | 3) −31.9 | |||
| D5-2 | Single site | 2.30 (±0.01) | 763.4 (±27.5) | −1459.1 (±1.1) | −382.7 | −35.1 |
| HK 496-516 | Single site | 2.56 (±0.02) | 1612.9 (±90.3) | −164.0 (±1.8) | −439.6 | −33.2 |
| gC1qR construct . | D5 construct . | N . | KD1 (nM) . | KD2 (nM) . | KD3 (nM) . | |
|---|---|---|---|---|---|---|
| WT | D5 | — | 1.9 (±0.1) | 64.9 (±1.9) | 1011.1 (±75.8) | |
| G2-5Ala | D5 | — | 1.2 (±0.1) | 33.4 (±5.7) | 724.6 (±178.3) | |
| DelG1 | D5 | — | 1.5 (±0.2) | 44.8 (±6.0) | 1283.7 (±238.8) | |
| G1-5Ala | D5 | — | 3.4 (±0.4) | 120.3 (±11.5) | 1083.4 (±176.6) | |
| DelG3 | D5 | No fit | ||||
| WT | D5-1 | — | 73.0 (±4.0) | 1579.8 (±145.3) | 2673.8 (±257.7) | |
| G2-5Ala | D5-1 | — | 131.1 (±43.0) | 1084.6 (±517.4) | 3663.0 (±369.1) | |
| DelG1 | D5-1 | — | 135.8 (±37.3) | 1228.5 (±407.9) | 3690.0 (±638.4) | |
| G1-5Ala | D5-1 | — | 120.8 (±55.4) | 680.3 (±420.0) | 1557.3 (±369.1) | |
| DelG3 | D5-1 | No fit | ||||
| WT | D5-2 | 2.3 (±0.01) | 763.4 (±27.5) | |||
| G2-5Ala | D5-2 | 2.1 (±0.02) | 885.0 (±31.9) | |||
| DelG1 | D5-2 | 2.3 (±0.03) | 724.6 (±50.0) | |||
| G1-5Ala | D5-2 | 2.3 (±0.02) | 800.0 (±45.6) | |||
| DelG3 | D5-2 | 0.5 (±0.02) | 2032.5 (±233.7) | |||
| gC1qR construct . | D5 construct . | N . | KD1 (nM) . | KD2 (nM) . | KD3 (nM) . | |
|---|---|---|---|---|---|---|
| WT | D5 | — | 1.9 (±0.1) | 64.9 (±1.9) | 1011.1 (±75.8) | |
| G2-5Ala | D5 | — | 1.2 (±0.1) | 33.4 (±5.7) | 724.6 (±178.3) | |
| DelG1 | D5 | — | 1.5 (±0.2) | 44.8 (±6.0) | 1283.7 (±238.8) | |
| G1-5Ala | D5 | — | 3.4 (±0.4) | 120.3 (±11.5) | 1083.4 (±176.6) | |
| DelG3 | D5 | No fit | ||||
| WT | D5-1 | — | 73.0 (±4.0) | 1579.8 (±145.3) | 2673.8 (±257.7) | |
| G2-5Ala | D5-1 | — | 131.1 (±43.0) | 1084.6 (±517.4) | 3663.0 (±369.1) | |
| DelG1 | D5-1 | — | 135.8 (±37.3) | 1228.5 (±407.9) | 3690.0 (±638.4) | |
| G1-5Ala | D5-1 | — | 120.8 (±55.4) | 680.3 (±420.0) | 1557.3 (±369.1) | |
| DelG3 | D5-1 | No fit | ||||
| WT | D5-2 | 2.3 (±0.01) | 763.4 (±27.5) | |||
| G2-5Ala | D5-2 | 2.1 (±0.02) | 885.0 (±31.9) | |||
| DelG1 | D5-2 | 2.3 (±0.03) | 724.6 (±50.0) | |||
| G1-5Ala | D5-2 | 2.3 (±0.02) | 800.0 (±45.6) | |||
| DelG3 | D5-2 | 0.5 (±0.02) | 2032.5 (±233.7) | |||
ITC-derived thermodynamic properties for the binding of HKD5, D5-1 and D5-2 with gC1qR, and individual KD values for the binding of HKD5 to the wt-gC1qR and gC1qR variant proteins. N values are not provided for the 3-site sequential binding fits, as the stoichiometry is fixed at 3. Values in parentheses is the error calculated based on the ITC curve fitting.