Kinetics of sodiumd-glucose cotransport in bovine intestinal brush border vesicles

Summary

Brush border membrane vesicles (BBMV) purified from steer jejunum were used to study the kinetics of sodiumd-glucose cotransport under voltage clamped, zero-trans conditions. When the initial rate of glucose transport (J _gluc) was measured over a wide range of glucose concentrations ([S]=0.01–20mm), curvature of the Woolf-Augustinsson-Hofstee plots was seen, compatible with a diffusional and one major, high capacity (maximal transport rateJ _max=5.8–8.8 nmol/mg·min) saturable system. Further studies indicated that changes incis [Na] altered theK _t , but not theJ _max, suggesting the presence of a rapid-equilibrium, ordered bireactant system with sodium adding first.Trans sodium inhibitedJ _gluc hyperbolically. KCl-valinomycin diffusion potentials, inner membrane face positive, loweredJ _gluc, while potentials of the opposite polarity raiseJ _gluc. At low glucose concentrations ([S]<0.05mm), a second, minor, high affinity transport system was indicated. Further evidence for this second saturable system was provided by sodium activation curves, which were hyperbolic when [S]=0.5mm, but were sigmoidal when [S]=.0.01mm. Simultaneous fluxes of²²Na and [³H]glucose at 1mm glucose and 30mm NaCl yielded a cotransport-dependent flux ratio of 2∶1 sodium/glucose, suggestive of 1∶1 (Na/glucose) high capacity, low affinity system and a ∼3∶1 (Na/glucose) high affinity, low capacity system. Kinetic experiments with rabbit jejunal brush borders revealed two major Na-dependent saturable systems. Extravesicular (cis) Na changed theK _t , but not theJ _max of the major system.