Membrane function in differentiating skeletal muscle cells: I. Kinetic analysis of amino acid transport |
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Authors: | Barbara Kay Grove Frank E. Stockdale |
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Affiliation: | 1. Department of Medicine, Stanford University, Stanford, California 94305 USA;2. Department of Biological Sciences, Stanford University, Stanford, California 94305 USA |
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Abstract: | Primary cultures of mononucleated myoblasts from 12-day-old chick embryos have a twofold higher rate of α-aminoisobutyric acid (AIB) transport before fusion occurs to form multinucleated myotubes. Several lines of evidence indicate that the uptake of AIB observed in both myoblasts and myotubes is primarily carrier-mediated by a membrane transport system. Increasing the temperature from 24 to 37°C results in a threefold increase in the rate of AIB uptake; both methionine and glycine inhibit AIB uptake by more than 85%; and 2,4-dinitrophenol inhibits AIB uptake by approximately 50%. In addition, the energies of activation (14.5 and 14.0 kcal/mole for myoblasts and myotubes, respectively) are characteristic of carrier-mediated transport. Resolution of AIB uptake into a saturable, carrier-mediated component and a nonsaturable, diffusion component shows that at concentrations of AIB≤1.5 mM over 97% of total AIB uptake is carriermediated in both myoblasts and myotubes. Kinetic analysis of carrier-mediated AIB uptake indicates that myoblasts and myotube membrane carriers have the same affinity for AIB (Km values = 1.73 and 1.31 mM, respectively). However, the Vmax for myoblasts is 23.7 nmole/mg/min while myotubes have a Vmax of 12.6 nmole/mg/min. The twofold difference in Vmax is shown to be due to a twofold difference in the quantity of membrane transport sites per milligram of protein. |
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