Marta M, Patamia M, Colella A, Sacchi S, Pomponi M, Kovacs K M, Lydersen C, Giardina B
Istituto di Chimica e Chimica Clinica, Centro CNR per la Chimica dei Recettori, UCSC, Facoltà di Medicina, Roma, Italy.
Biochemistry. 1998 Oct 6;37(40):14024-9. doi: 10.1021/bi981157v.
It is generally believed that bovine hemoglobin (BvHb) interacts weakly with 2,3-diphosphoglycerate (2,3-DPG) in a chloride-free media and not at all in the presence of physiological concentrations of chloride (100 mM). This lack of interaction has raised several questions at both structural and evolutionary levels. Results obtained in this study via 31P nuclear magnetic resonance (NMR) show that, even in the presence of 100 mM chloride ions, 2,3-DPG does, in fact, interact with bovine deoxy-Hb. This spectroscopic observation has been confirmed by oxygen binding experiments, which have also shown that, under certain conditions, chloride and 2,3-DPG may display a synergistic effect in modifying the oxygen affinity of bovine hemoglobin. It could be that this synergistic effect has its structural basis in a conformational modification induced by 2,3-DPG, possibly causing extra chloride anions to approach the positive charges which constitute the anion binding site. Another possibility, not necessarily an alternative, is the additional chloride binding site recently identified [Fronticelli, C., Sanna, M. T., Perez-Alvarado, G. C., Karavitis, M., Lu, A.-L., and Brinnigar, W. S. (1995) J. Biol. Chem 270, 30588-30592] involving lysine beta76 that in bovine Hb substitutes for the alanine residue present in human hemoglobin. All of these findings are in agreement with the very low enthalpy of oxygenation that characterizes bovine Hb when both chloride and 2,3-DPG are present in concomitance. The results reported here clearly show that bovine hemoglobin does react with 2, 3-DPG and is functionally affected by this organic phosphate. Hence, the very low intraerythrocytic concentration of 2,3-DPG (0.5 mM) in adult bovine red blood cells is the result of metabolic adaptation which cannot be explained solely by the different amino acid sequence at the level of the 2,3-DPG binding site.
一般认为，在无氯化物介质中，牛血红蛋白（BvHb）与2,3-二磷酸甘油酸（2,3-DPG）的相互作用较弱，而在生理浓度的氯化物（100 mM）存在时则完全不相互作用。这种缺乏相互作用在结构和进化层面都引发了几个问题。本研究通过31P核磁共振（NMR）获得的结果表明，即使在存在100 mM氯离子的情况下，2,3-DPG实际上确实与牛脱氧血红蛋白相互作用。这一光谱学观察结果已通过氧结合实验得到证实，该实验还表明，在某些条件下，氯离子和2,3-DPG在改变牛血红蛋白的氧亲和力方面可能表现出协同效应。可能这种协同效应在结构上基于2,3-DPG诱导的构象修饰，可能导致额外的氯离子阴离子接近构成阴离子结合位点的正电荷。另一种可能性，不一定是替代方案，是最近发现的[Fronticelli, C., Sanna, M. T., Perez-Alvarado, G. C., Karavitis, M., Lu, A.-L., and Brinnigar, W. S. (1995) J. Biol. Chem 270, 30588 - 30592]涉及赖氨酸β76的额外氯离子结合位点，在牛血红蛋白中该位点替代了人类血红蛋白中存在的丙氨酸残基。所有这些发现都与当氯离子和2,3-DPG同时存在时表征牛血红蛋白的非常低的氧合焓一致。这里报道的结果清楚地表明，牛血红蛋白确实与2,3-DPG反应，并在功能上受到这种有机磷酸盐的影响。因此，成年牛红细胞中2,3-DPG的极低细胞内浓度（0.5 mM）是代谢适应的结果，这不能仅通过2,3-DPG结合位点水平上不同的氨基酸序列来解释。
