Table 2.
CFP fluorescent lifetimes and CFP/YFP Förster efficiency in dual-tagged GluA2-6Y-10C receptors in the presence of glutamate and CTZ
| Receptor | I6 insert | I10 insert | Ligand | τCFP,† ns | n | E,‡ % |
| GluA2 | CFP | YFP | None | 3.05 ± 0.02 | 31 | 13.4 |
| GluA2 | CFP | amber | None | 3.52 ± 0.03 | 17 | — |
| GluA2 | CFP | — | None | 3.46 ± 0.03 | 8 | — |
| GluA2-S754Q | CFP | YFP | None | 2.99 ± 0.01 | 11 | 16.8 |
| GluA2-S754Q | CFP | amber | None | 3.56 ± 0.01 | 11 | — |
| GluA2 | CFP | YFP | GLU | 3.02 + 0.03 | 15 | 12.7 |
| GluA2 | CFP | amber | GLU | 3.47 + 0.03 | 11 | — |
| GluA2 | CFP | — | GLU | 3.41 + 0.03 | 8 | — |
| GluA2-S754Q | CFP | YFP | GLU | 2.95 + 0.04 | 10 | 18.2 |
| GluA2-S754Q | CFP | amber | GLU | 3.60 + 0.01 | 10 | — |
| GluA2 | CFP | YFP | CTZ | 3.17 + 0.02* | 9 | 8.0 |
| GluA2 | CFP | amber | CTZ | 3.44 + 0.03 | 9 | — |
| GluA2 | CFP | — | CTZ | 3.41 + 0.02 | 6 | — |
| GluA2-S754Q | CFP | YFP | CTZ | 2.99 + 0.03 | 11 | 16.8 |
| GluA2-S754Q | CFP | amber | CTZ | 3.59 + 0.01 | 10 | — |
| GluA2 | CFP | YFP | GLU + CTZ | 3.14 ± 0.01* | 10 | 9.0 |
| GluA2 | CFP | amber | GLU + CTZ | 3.45 ± 0.02 | 10 | — |
| GluA2 | CFP | — | GLU + CTZ | 3.40 ± 0.02 | 8 | — |
| GluA2-S754Q | CFP | YFP | GLU + CTZ | 2.98 + 0.03 | 13 | 17.9 |
| GluA2-S754Q | CFP | amber | GLU + CTZ | 3.63 + 0.03 | 10 | — |
GLU, glutamate.
*
P < 0.05 compared with same construct with no ligand.
†
Fluorescence lifetime for CFP in the presence of GLU (1,000 μM) and/or CTZ (25 μM) was obtained by FLIM analysis as described in Materials and Methods. Data represent the mean ± SEM of the average lifetime observed in n different cells.
‡
FRET efficiency was calculated as described in Materials and Methods from CFP lifetimes obtained for GluA2-6Y-10C and the corresponding donor-only amber construct.