Ginsburg H
Biochim Biophys Acta. 1978 Jan 4;506(1):119-35. doi: 10.1016/0005-2736(78)90439-x.
The kinetic properties of the mediated transport of galactose in human erythrocytes are investigated at 20 degrees C. Different methodological procedures are used to acquire a complete kinetic description of the system. Under zero-trans conditions the uptake of galactose is mediated by two distinctly different carriers (defined as alpha and beta) having significantly different Mic;aelis parameters: alpha K = 12.7 mM and beta K = 81.5 mM, but similar maximal velocities, approx. 40 nM.min-1. The zero-trans efflux procedure reveals apparently one single carrier with K = 74.4 mM and V = 241 mM.min-1. Under equilibrium-exchange conditions the galactose transport is mediated apparently by a single site with K = 146 mM and V = 521 mM.min-1. The data for the alpha-carrier are analyzed in terms of the simple carrier model as formulated by Lieb and Stein (Biochim. Biophys. Acta (1974) 373, 178). Application of several rejection criteria for the simple carrier failed to indicate lack of fitness of the alpha-carrier to a simple asymmetric carrier. From the analysis of the kinetic data it is inferred that the transport of galactose across the human erythrocyte membrane is mediated by two simple asymmetric carriers operating in antiparallel fashion. Using this model and the data of zero-trans and equilibrium-exchange, it is shown that the predicted half-saturation constants for both uptake and efflux in infinite-cis conditions fully agree with the experimentally derived values. Further analysis of the kinetic data indicate that the translocation of the unloaded alpha-carrier is the rate-limiting step in galactose uptake. Under equilibrium-exchange conditions the unloaded carrier is asymmetrically distributed across the membrane so that its concentration is 8 times higher on the inner side of the membrane. Using the value of 3.3.10(5) hexose carriers per cell, the turnover number of galactose exchange is 6.5.10(4) molecules/carriers per min.
在20摄氏度下研究了半乳糖在人红细胞中介导转运的动力学特性。采用了不同的方法程序来获得该系统完整的动力学描述。在零转运条件下，半乳糖的摄取由两种明显不同的载体（定义为α和β）介导，它们具有显著不同的米氏参数：αK = 12.7 mM，βK = 81.5 mM，但最大速度相似，约为40 nM·min⁻¹。零转运外排程序显示明显只有一种载体，K = 74.4 mM，V = 241 mM·min⁻¹。在平衡交换条件下，半乳糖转运显然由一个单一位点介导，K = 146 mM，V = 521 mM·min⁻¹。根据Lieb和Stein（《生物化学与生物物理学报》（1974年）373卷，178页）提出的简单载体模型对α载体的数据进行了分析。对简单载体应用多种排除标准未能表明α载体不适合简单的不对称载体。从动力学数据分析推断，半乳糖跨人红细胞膜的转运由两种以反平行方式运作的简单不对称载体介导。使用该模型以及零转运和平衡交换的数据表明，在无限顺式条件下摄取和外排的预测半饱和常数与实验得出的值完全一致。对动力学数据的进一步分析表明，空载α载体的转位是半乳糖摄取中的限速步骤。在平衡交换条件下，空载载体不对称地分布在膜上，使得其在膜内侧的浓度高8倍。利用每个细胞有3.3×10⁵个己糖载体这一数值，半乳糖交换的周转数为6.5×10⁴个分子/载体每分钟。
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