Himmelreich U, Chapman B E, Kuchel P W
Department of Biochemistry, University of Sydney, NSW, Australia.
Eur Biophys J. 1999;28(2):158-65. doi: 10.1007/s002490050195.
The rate of the rapid exchange of formate mediated by band 3 in human erythrocytes, under equilibrium exchange conditions, was measured by using a T1 relaxation method with 13C-labelled formate and 13C NMR, and a pulsed field-gradient spin-echo (PFGSE) method using 1H NMR. The former analysis was based on large differences in T1 between the inside and the outside of the cells brought about by added Mn2+; the latter was based on large differences in the apparent diffusion coefficient inside and outside the cells. There was close agreement in the estimates of the membrane permeabilities made using both methods, suggesting a lack of interference of the exchange process by Mn2+. Regression analysis yielded estimates (under the specified conditions, including 37 degrees C) of Vmax of 3.5 +/- 0.3 x 10(-9) and 3.8 +/- 0.4 x 10(-9) mol cm-2 s-1, and K(m) of 9.8 +/- 0.2 and 8.1 +/- 0.2 mM, for the T1 and the PFGSE methods, respectively. These are new estimates made using methodology that has not previously been applied to measuring rapid (sub-second time scale) formate exchange in cells.
在平衡交换条件下，利用含(^{13}C)标记甲酸盐的(T1)弛豫方法和(^{13}C)核磁共振以及利用(^{1}H)核磁共振的脉冲场梯度自旋回波（PFGSE）方法，测量了人红细胞中由带3介导的甲酸盐快速交换速率。前一种分析基于添加(Mn^{2+})导致细胞内外(T1)的巨大差异；后一种分析基于细胞内外表观扩散系数的巨大差异。两种方法得出的膜渗透率估计值非常一致，这表明(Mn^{2+})对交换过程没有干扰。回归分析得出（在指定条件下，包括37摄氏度），对于(T1)方法和PFGSE方法，(Vmax)分别为(3.5\pm0.3\times10^{-9})和(3.8\pm0.4\times10^{-9}) (mol\ cm^{-2}\ s^{-1})，(K(m))分别为(9.8\pm0.2)和(8.1\pm0.2) (mM)。这些是使用以前未应用于测量细胞中快速（亚秒时间尺度）甲酸盐交换的方法得出的新估计值。
