AJP - Gastrointestinal and Liver Physiology, Vol 259, Issue 5 865-G872, Copyright © 1990 by American Physiological Society
Transport of biotin in basolateral membrane vesicles of rat liver
H. M. Said, S. Korchid, D. W. Horne and M. Howard
Department of Medicine, University of California School of Medicine, Irvine 92717.
We examined biotin transport across the basolateral membrane (BLM) of rat
liver using BLM vesicles (BLMV) technique. The purity and suitability for
transport studies of liver BLMV were demonstrated by morphological
(electron microscopy), enzymatic, and functional criteria. Orientation of
liver BLMV was determined by freeze-fracture electron microscopy and by
[3H]ouabain binding methodology and was found to be 65.3-69.7% in the
right-side-out orientation. Uptake of biotin by liver BLMV was found by
osmolarity and temperature studies to be mostly the result of transport of
the substrate into an active intravesicular space with little binding to
membrane surfaces. Transport of biotin was found to be Na+ gradient
dependent with a distinct "over-shoot" phenomenon. Initial rate of
transport of biotin as a function of concentration was found to include a
saturable component in the presence of a Na+ gradient (out greater than in)
but was linear and lower in the presence of a K+ gradient (no Na+). Kinetic
parameters of the saturable Na+ gradient-dependent transport process were
0.39 microM and 1,807 fmol.mg protein-1.20 s-1 for the apparent Km and
Vmax, respectively. In the presence, but not the absence, of a Na+ gradient
(out greater than in), the addition of structural analogues to the
incubation medium caused significant inhibition in the transport of 0.079
microM [3H]biotin. Induction, with the use of valinomycin and an inwardly
directed K+ gradient, of a relatively positive intravesicular space caused
significant inhibition in the initial rate of biotin transport.(ABSTRACT
TRUNCATED AT 250 WORDS)
