Figure 3.

Expression of recombinant hSGLT1 in E. coli. (A) Sugar uptake (50 μM α-MDG) by right-side-out vesicles of E. coli BL21 producing hSGLT1ΔN is stimulated by sodium (Na⁺, 25 mM) and inhibited by phlorizin (Phlz, 10 μM). No Na⁺-dependent, phlorizin-sensitive uptakes were observed with vesicles of BL21 harboring plasmid pT7–5 as a control (Co). For the control vesicles, there was no significant difference in sugar uptake in the presence or absence of Na⁺ (39 ± 12 nmol α-MDG × mg protein⁻¹ × min⁻¹), which was not affected by the addition of phlorizin. (B) Western blot analysis of the membrane fraction of E. coli B21 producing hSGLT1ΔN. No signal was detected in other cell fractions. The apparent molecular mass of hSGLT1 corresponds to that of the nonglycosylated transporter in Xenopus oocytes (compare Fig. 2).