Table 1.
Quantification of STxB retrograde transport and Gb3 expression
| Cell types | Glycosylation (%) | Gb3 (μg/106 cells) | Binding sites (×106/cell) | KD (nM) |
|---|---|---|---|---|
| HeLa | 7.3 ± 1.4 (10) | 0.94 ± 0.16 (9) | 40 | 19 |
| HeLa +PPMP | 6.3 ± 1.7 (7) | 0.25 ± 0.05 (5) | 1.2 ± 0.6 (4) | 19 ± 7 (4) |
| Macrophages | background | 0.04 ± 0.02 (4) | 0.4 ± 0.2 (4) | 85 ± 30 (4) |
| Macrophage +LPS | 1.1 ± 0.5 (3) | 0.27 ± 0.1 (4) | 2.8 ± 1.2 (4) | 21 ± 3 (4) |
| Monocytes | background | 0.06 ± 0.03 (3) | n.d. | n.d. |
| Monocytes +LPS | 1.5 ± 0.5 (3) | 0.27 ± 0.2 (4) | n.d. | n.d. |
| imDC | background | 0.04 (2) | n.d. | n.d. |
n.d., not determined.
STxB-Glyc-KDEL was bound to the indicated cells on ice and after washings, the cells were incubated for 4 h at 37°C. Arrival in the ER was quantified through the detection of the protein's glycosylation (percentage of glycosylated, cell-associated protein). Gb3 was quantified on TLC plates after extraction of neutral glycolipids and detection by overlay. Binding sites and the apparent affinity of STxB-Glyc-KDEL for the indicated cells were determined by Scatchard analysis. Data are given as means ± SEM (number of experiments in parentheses).