Indiveri C, Tonazzi A, Palmieri F
Department of Pharmaco-Biology, University of Bari, Italy.
Biochim Biophys Acta. 1991 Oct 14;1069(1):110-6. doi: 10.1016/0005-2736(91)90110-t.
The carnitine carrier from rat liver mitochondria was purified by chromatography on hydroxyapatite and celite and reconstituted in egg yolk phospholipid vesicles by adsorbing the detergent on polystyrene beads. In the reconstituted system, in addition to the carnitine/carnitine exchange, the purified protein catalyzed a uni-directional transport (uniport) of carnitine measured as uptake into unloaded proteoliposomes as well as efflux from prelabelled proteoliposomes. In both cases the reaction followed a first-order kinetics with a rate constant of 0.023-0.026 min-1. Besides carnitine, also acylcarnitines were transported in the uniport mode. N-Ethylmaleimide inhibited the uni-directional transport of carnitine completely. The uniport of carnitine is not influenced by the delta pH and the electric gradient across the membrane. The activation energy for uniport was 115 kJ/mol and the half-saturation constant on the external side of the proteoliposomes was 0.53 mM. The maximal rate of the uniport at 25 degrees C was 0.2 mumol/min per mg protein, i.e. about 10 times lower than that of the reconstituted carnitine transport in exchange mode.
通过羟基磷灰石和硅藻土色谱法对大鼠肝脏线粒体中的肉碱载体进行纯化，并通过将去污剂吸附在聚苯乙烯珠粒上，将其重构成蛋黄磷脂囊泡。在重构系统中，除了肉碱/肉碱交换外，纯化的蛋白质还催化了肉碱的单向转运（单向运输），其通过测定未负载的蛋白脂质体的摄取以及预标记的蛋白脂质体的流出进行测量。在这两种情况下，反应均遵循一级动力学，速率常数为0.023 - 0.026 min⁻¹。除了肉碱外，酰基肉碱也以单向运输模式进行转运。N - 乙基马来酰亚胺完全抑制了肉碱的单向转运。肉碱的单向运输不受跨膜ΔpH和电势梯度的影响。单向运输的活化能为115 kJ/mol，蛋白脂质体外侧的半饱和常数为0.53 mM。在25℃下单向运输的最大速率为每毫克蛋白质0.2 μmol/min，即比以交换模式重构的肉碱转运速率低约10倍。
