Hexose transport in L6 muscle cells. Kinetic properties and the number of [3H] cytochalasin B binding sites
A Klip, WJ Logan, G Li - Biochimica et Biophysica Acta (BBA) …, 1982 - Elsevier
A Klip, WJ Logan, G Li
Biochimica et Biophysica Acta (BBA)-Biomembranes, 1982•ElsevierMyoblasts in culture (L6 cell line) were used as an in vitro model system, to study the kinetic
and pharmacological properties of hexose transport in skeletal muscle tissue.(2) Uptake of 2-
deoxy-d-[3 H] glucose into L6 cells grown in monolayer culture was judged rate limiting
since:(a) The time course of sugar uptake extrapolated to zero,(b) a parallel inhibition of
hexose uptake and phosphorylation was caused by cytochalasin B, and (c) very little
backflow of the hexose was detected.(3) Uptake of 2-deoxy-d-[3 H] glucose by cells in …
and pharmacological properties of hexose transport in skeletal muscle tissue.(2) Uptake of 2-
deoxy-d-[3 H] glucose into L6 cells grown in monolayer culture was judged rate limiting
since:(a) The time course of sugar uptake extrapolated to zero,(b) a parallel inhibition of
hexose uptake and phosphorylation was caused by cytochalasin B, and (c) very little
backflow of the hexose was detected.(3) Uptake of 2-deoxy-d-[3 H] glucose by cells in …
Myoblasts in culture (L6 cell line) were used as an in vitro model system, to study the kinetic and pharmacological properties of hexose transport in skeletal muscle tissue.(2) Uptake of 2-deoxy-d-[3 H] glucose into L6 cells grown in monolayer culture was judged rate limiting since:(a) The time course of sugar uptake extrapolated to zero,(b) a parallel inhibition of hexose uptake and phosphorylation was caused by cytochalasin B, and (c) very little backflow of the hexose was detected.(3) Uptake of 2-deoxy-d-[3 H] glucose by cells in monolayers was linear for at least 20 min and it was stimulated by countertransport. The K t value was 0.83 mM. Cytochalasin B inhibited uptake non-competitively, and half maximal inhibition was achieved at 0.3 μM. Cytochalasin E (up to 5 μM) did not affect 2-deoxy-d-[3 H] glucose uptake.(4) L6 myoblasts, detached by trypsinization, retained the hexose transport activity. K t in detached cells was 0.96 mM. V was 3.2 nmol/min per mg protein, and half maximal inhibition was observed with 0.25 μM cytochalasin B.(5)[3 H] Cytochalasin B binding to detached cells showed saturable and non-saturable components. The former could be further separated into cytochalasin E-sensitive binding (probably associated to cytoskeletal proteins) and cytochalasin E-insensitive binding, a fraction of which was inhibited by d-glucose. The d-glucose sensitive sites amount to 16.3 pmol/mg protein, and showed a K d of 0.49 μM, which is in close agreement with the K i of cytochalasin B inhibition of hexose uptake. These sites probably are equivalent to the hexose carrier molecules, and are present at a density of 6.8· 10 6 sites/cell.
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