Fig. 4: Continuous wave FCS hints at additional slow rotation.
a Exemplary fullFCS measurement from a CHO-K1 cell transfected with N-terminal-EGFP-β2-AR. For the data sliced in 5 s pieces a difference in amplitude between the CCFVV-VH (black) and ACFVH (light green) compared ACFVV (green) can be seen. The data was globally fitted with Eq. (10). Weighted residues are shown on top. b The absolute difference in amplitude between ACFVV and ACFVH calculated from the fit parameter (Eq. (12)). c For the ACFVH-curves, an increase in amplitude of τrot leads to decreasing amount of the two τR in the µs range. Both τR1 and τR2 are positively correlated. Each color represents the measurement from one cell (n = 7). τrot = 99 ± 52 ns, τR1 = 1.90 ± 0.45 µs, τR2 = 128 ± 39 µs. Data are mean ± s.d. d An increased amount of slow, membrane diffusion, xslow, is related to a decreasing amplitude of τrot. The color code is same as in (a). Results for CCFVV–VH and ACFVV and for the N-terminal-SNAP-β2AR conjugated to AF488 are summarized in Supplementary Figs. 16c–f and S17.
