L-Leucine transport in human red blood cells: a detailed kinetic analysis

The kinetic properties of L-leucine transport across the human red blood cell membrane was analyzed according to the simple pore and carrier theory of Lieb and Stein (Biochim. Biophys. Acta, 1974, 373: 165-177 and 178-196) at 25 degrees C, pH 7.4. Several methods were used in order to obtain a thorough kinetic description of L-leucine transport. A rejection of the simple pore model was suggested from the result of zero-trans influx and zero-trans and equilibrium-exchange efflux experiments. Several predictions from the simple carrier model, based on the requirement of consistency among different kinetic parameters, were tested in infinite experiments, i.e. experiments performed at a high concentration of substrate at one of the faces of the membrane. The simple pore model was rejected, but no crucial evidence against a simple carrier model, which displays symmetric properties at 25 degrees C, was found in the concentration range considered (0.002-68 mM). The relative magnitudes of the rate constants of the translocation process are discussed, and it is concluded (a) that both the dissociation and translocation of carrier-complex is faster than the translocation of the empty carrier, (b) that no translocation step is rate determining, and (c) that the carrier-complex is equally distributed across the membrane at equilibrium. The present work provides a unique example of a carrier-mediated transport mechanism which displays symmetric properties. L-leucine transport in red blood cells may be a convenient system for studying molecular mechanisms of facilitated transport.