Fig. 3: Time course of binary-FRET of CaMKII activation and NR2B association.

Changes in the in vitro mVenus fluorescence lifetime (a) or steady-state anisotropy (b) when V-CaMKII was activated alone (green circles), or in the presence of mCherry-tagged NR2B binding fragment (red triangles), mUranus-tagged NR2B binding fragment (gray squares), or with mCherry-tagged mutant NR2B(L1303A) binding fragment (yellow diamonds). Each point is mean ± SD, n = 4 independent samples with two replicates per sample. Representative anisotropy and lifetime decay traces are shown in Supplementary Fig. 2. Live cell binary-FRET lifetime (c and e) and anisotropy (d and f) images of HEK cells co-expressing V-CaMKII, mCh-NR2B-binding fragment, and calmodulin before (c and d) and after (e and f) activation with ionomycin. Anisotropy color bars ranged from 0.3 (dark red) to 0.5 (dark purple). Lifetime color bars are in picoseconds and range from 0 ps (dark red) to 3500 ps (dark purple). Source data are provided as a Source Data file.