Kd values determined for GluA2 constructs by different techniques
| Technique | Temperature | GluA2S | GnTI− | EndoH | GluA2L | GluA2S-FAM | EndoH-FAM | EndoH-DyLight |
| SV | 10°C | 6.0a [2.1–22] | 14.9b [7.6–47] | |||||
| 20°C | 8.3 [2.0–22] | 11 [0.8–43] | 12c [7.0–30] | 5.5 [2.8–43] | ||||
| 8.3 [5.1–27] | ||||||||
| 10 [3.0–20] | ||||||||
| FDS-SV | 20°C | 13d [0.76–203] | ||||||
| 2.5e [0.4–7.8] | ||||||||
| 10f [3.6–19] | ||||||||
| SE | 4°C | 160 [83–268] | ||||||
| 284 [159–461] | ||||||||
| 30g [ND–260] | ||||||||
| 10°C | 13h [0.14–50] | |||||||
| 17 [ND–74] | ||||||||
| 244h [46–643] | ||||||||
| FAI | 20°C | 10.8 [2.4–29] | ||||||
| 11.3 [ND–68] | ||||||||
| 8.43 [ND–50.8] |
| Technique | Temperature | GluA2S | GnTI− | EndoH | GluA2L | GluA2S-FAM | EndoH-FAM | EndoH-DyLight |
| SV | 10°C | 6.0a [2.1–22] | 14.9b [7.6–47] | |||||
| 20°C | 8.3 [2.0–22] | 11 [0.8–43] | 12c [7.0–30] | 5.5 [2.8–43] | ||||
| 8.3 [5.1–27] | ||||||||
| 10 [3.0–20] | ||||||||
| FDS-SV | 20°C | 13d [0.76–203] | ||||||
| 2.5e [0.4–7.8] | ||||||||
| 10f [3.6–19] | ||||||||
| SE | 4°C | 160 [83–268] | ||||||
| 284 [159–461] | ||||||||
| 30g [ND–260] | ||||||||
| 10°C | 13h [0.14–50] | |||||||
| 17 [ND–74] | ||||||||
| 244h [46–643] | ||||||||
| FAI | 20°C | 10.8 [2.4–29] | ||||||
| 11.3 [ND–68] | ||||||||
| 8.43 [ND–50.8] |
Kd values are reported in nanomolar units. Errors represent the 95% CI using an automated surface projection method, unless indicated otherwise. 10 independent AUC experiments were performed for GluA2S digested with EndoH, as indicated by replicate entries for the mean and 95% CI. SV, sedimentation velocity with absorbance and interference optics; FDS-SV, sedimentation velocity with fluorescence detection optics; SE, sedimentation equilibrium with absorbance optics; FAI, fluorescence anisotropy.
68.3% CI.
The data obtained after FAM labeling of GluA2 led to the highest best-fit value, but the labeling does not significantly affect binding within the 95% CI of this assay.
Global analysis of the absorbance data at 230 nm shown in Fig. 3 D, with additional data acquired from the same experiment at 210 nm shown in Fig. S2 (top right).
Analysis of FDS-SV data only without hydrodynamic constraints.
Analysis of FDS-SV data only with hydrodynamic constraints.
Analysis of FDS-SV data with one additional single high concentration data point measured by absorbance at 488 nm.
Analysis with low loading concentration and 210-nm detection.
Analysis with oil layer to increase pressure.