Figure 2.

FcγRIIB-T232 exhibits significantly slower Brownian diffusion than FcγRIIB-I232 in human primary B cells and monocytes. (A and B) Comparison of the Brownian diffusion of FcγRIIB-I232 or FcγRIIB-T232 molecules from either human primary B cells (A) or primary monocytes (B). (C) Comparison of the Brownian diffusion of FcγRIIB and IgM-BCRs molecules in human primary B cells. (A–C) Cells were placed on coverslips tethering anti–MHC-I antibodies in TIRF microscopy imaging. Given are a series of mathematical comparisons in CPD plots (left), MSD plots (middle), and scatter plots (right; bars represent the median value). The p-value in each CDP plots is <0.0001 in Kolmogorov-Smirnov tests. The results shown are representative of one of at least three independent experiments. (D) Monte Carlo simulation of receptors’ binding time. The times for first binding between 625 receptors diffusing at the indicated diffusion rate and 5 random distributed immobile ICs that were confined in a 2.5 × 2.5–µm square were calculated. Error bars represent mean ± SEM from 400× simulation per group that were pooled from at least two independent experiments. (E) In this illustrative diagram, the TM domain is represented as a rectangle colored in gray, whereas the plasma membrane is colored in cyan with the borders indicated by black solid lines. The red solid bars represent the projections of the TM helices within the plane of the plasma membrane, which equal h × tan(α), where h is the thickness of the membrane (denoted as black dashed lines) and α is the tilting angle of a TM helix. The cross-sectional area shall be proportional to tan²(α). The FcγRIIB-I232 TM helix prefers the tilting orientation of 13–18°, whereas the FcγRIIB-T232 TM helix is the most stable at the tilting angle of 35–40° (Fig. 6 J). Therefore, the ratio of cross-sectional area from the FcγRIIB-T232 TM helix to that from FcγRIIB-I232 shall fall in the interval between [tan(35°)/tan(18°)]² and [tan(40°)/tan(13°)]², or [4.6, 13.2].