Gasbjerg P K, Brahm J
Department of General Physiology and Biophysics, Panum Institute, University of Copenhagen, Denmark.
Biochim Biophys Acta. 1991 Feb 11;1062(1):83-93. doi: 10.1016/0005-2736(91)90338-9.
D-[14C]Glucose self exchange and unidirectional efflux from human red blood cells were studied at 20 degrees C (pH 7.2) by means of the Millipore-Swinnex filtering technique whose time resolution is greater than 1 s and the continuous flow-tube method with a time resolution of greater than 2 ms. The unidirectional efflux data were analyzed using both the method of initial rates and the integrated rate equation. Simple Michaelis-Menten kinetics apply to the results obtained under both experimental conditions. In self-exchange mode, the half-saturation constant, K1/2ex, was 10 (S.E. +/- 1) mM. In unidirectional efflux mode K1/2ue was 6.6 (S.E. +/- 0.5) mM (initial rates) or by the method of integrated rates 7.7 mM, with a range of 2.7-12.1 mM, K1/2ue increasing with an increased initial intracellular glucose concentration. Our results of K1/2ex oppose previous published values of 32 mM for self exchange (Eilam and Stein (1972) Biochim. Biophys. Acta 266, 161-173) and 25 mM for unidirectional efflux (Karlish et al. (1972) Biochim. Biophys. Acta 255, 126-132) that have been used extensively in kinetic considerations of glucose transport models. Under self-exchange conditions Jmaxex was 1.8 x 10(-10) mol cm-2s-1, and in unidirectional efflux mode Jmaxue was 8.3 x 10(-11) mol cm-2s-1 (initial rates) and 8.6 x 10(-11) mol cm-2s-1 (integrated rates). We suggest that the previous high values of Jmax and in particular K1/2 are due to the use of methods with insufficient time resolution. Our results indicate that the transport system is less asymmetric than was generally accepted, and that complicated transport models developed to account for the great difference between the determined K1/2 and J max values are redundant.
在20摄氏度（pH 7.2）下，采用时间分辨率大于1秒的密理博 - 斯温克斯过滤技术以及时间分辨率大于2毫秒的连续流管法，研究了D-[¹⁴C]葡萄糖在人红细胞中的自交换和单向流出。单向流出数据采用初始速率法和积分速率方程进行分析。简单的米氏动力学适用于在两种实验条件下获得的结果。在自交换模式下，半饱和常数K₁/₂ex为10（标准误差±1）毫摩尔。在单向流出模式下，K₁/₂ue为6.6（标准误差±0.5）毫摩尔（初始速率），或通过积分速率法为7.7毫摩尔，范围为2.7 - 12.1毫摩尔，K₁/₂ue随初始细胞内葡萄糖浓度的增加而增加。我们得到的K₁/₂ex结果与之前发表的自交换值32毫摩尔（Eilam和Stein（1972年），生物化学与生物物理学报266, 161 - 173）和单向流出值25毫摩尔（Karlish等人（1972年），生物化学与生物物理学报255, 126 - 132）相反，这些值在葡萄糖转运模型的动力学考虑中被广泛使用。在自交换条件下，Jmaxex为1.8×10⁻¹⁰摩尔·厘米⁻²·秒⁻¹，在单向流出模式下，Jmaxue为8.3×10⁻¹¹摩尔·厘米⁻²·秒⁻¹（初始速率）和8.6×10⁻¹¹摩尔·厘米⁻²·秒⁻¹（积分速率）。我们认为，之前较高的Jmax值，尤其是K₁/₂值，是由于使用了时间分辨率不足的方法。我们的结果表明，转运系统的不对称性比普遍认为的要小，并且为解释所确定的K₁/₂和Jmax值之间的巨大差异而开发的复杂转运模型是多余的。
