Weber R E
Zoophysiology Laboratory, Aarhus University, Denmark.
J Appl Physiol (1985). 1992 Apr;72(4):1611-5. doi: 10.1152/jappl.1992.72.4.1611.
The functional characteristics of hemoglobin (Hb) depend on oxygenation-linked proton and anion binding and thus on solvent buffer groups and ionic composition. This study compares the oxygenation properties of human Hb in ionic [tris(hydroxymethyl)aminomethane (Tris) and BisTris] buffers with those in zwitterionic N-2-hydroxy-ethylpiperazine-N'-2-ethanesulfonic acid (HEPES) buffer under strictly controlled chloride concentrations at different pH values, two temperatures, and in the absence and presence of the erythrocytic cofactor, 2,3-diphosphoglycerate (DPG). In contrast to earlier studies (carried out at the same or different chloride concentrations) it shows only small buffer effects that are manifested at low chloride concentration and high pH. These observations suggest chloride binding to the Tris buffers, which reduces the interaction with specific chloride binding sites in the Hb. The findings indicate that HEPES allows for more accurate assessment of Hb-oxygen affinity and its anion and temperature sensitivities than ionic buffers and advocates standard use of HEPES in studies on Hb function. Precise oxygen affinities of Hb dissolved in both buffers are defined under standard conditions.
血红蛋白（Hb）的功能特性取决于与氧合相关的质子和阴离子结合，因此取决于溶剂缓冲基团和离子组成。本研究比较了在不同pH值、两种温度下，在严格控制的氯化物浓度条件下，以及在不存在和存在红细胞辅因子2,3-二磷酸甘油酸（DPG）的情况下，人Hb在离子型[三（羟甲基）氨基甲烷（Tris）和双Tris]缓冲液与两性离子N-2-羟乙基哌嗪-N'-2-乙磺酸（HEPES）缓冲液中的氧合特性。与早期研究（在相同或不同氯化物浓度下进行）不同，本研究仅显示出在低氯化物浓度和高pH值时出现的微小缓冲效应。这些观察结果表明氯化物与Tris缓冲液结合，这减少了与Hb中特定氯化物结合位点的相互作用。研究结果表明，与离子型缓冲液相比，HEPES能更准确地评估Hb与氧的亲和力及其对阴离子和温度的敏感性，并提倡在Hb功能研究中标准使用HEPES。在标准条件下定义了溶解于两种缓冲液中的Hb的精确氧亲和力。
