Serbanescu R, Kiger L, Poyart C, Marden M C
University of Bucarest, Department of Biophysics, Magurele, Romania.
Biochim Biophys Acta. 1998 Jan 27;1363(1):79-84. doi: 10.1016/s0005-2728(97)00092-3.
A fluorescent analogue of diphosphoglycerate (DPG), hydroxy-pyrenetrisulfonate (HPT), was used as a probe of the allosteric equilibrium of methemoglobin. Like DPG, HPT binds, one per tetramer, with a higher affinity to deoxyHb than to oxyHb. Once bound, the HPT fluorescence is quenched by energy transfer to the hemes. HPT can thus serve as a probe of the conformational state of the hemoglobin tetramer: a higher quenching indicates a stronger binding and therefore, more of the deoxy conformation. Since HPT binds to the same site as DPG, it can be displaced by DPG in order to determine the fluorescence intensity of the free HPT under the same conditions, to correct for the inner filter effect. The high spin ferric ligands, such as water and F, showed less fluorescence (more of the deoxy state) than low spin cyano-metHb. The aquo-metHb samples showed a reversion to the oxyHb conformation above pH 7, as expected due to the acid-alkaline transition forming hydroxy-metHb. Effectors such as bezafibrate, which do not bind to the same site as DPG, show an increase in the deoxy-like characteristics.
二磷酸甘油酸（DPG）的荧光类似物羟基芘三磺酸盐（HPT）被用作高铁血红蛋白变构平衡的探针。与DPG一样，HPT以每个四聚体一个的比例结合，与脱氧血红蛋白的亲和力高于与氧合血红蛋白的亲和力。一旦结合，HPT荧光会通过能量转移到血红素而被淬灭。因此，HPT可以作为血红蛋白四聚体构象状态的探针：更高的淬灭表明更强的结合，因此更多的是脱氧构象。由于HPT与DPG结合在同一部位，它可以被DPG取代，以便在相同条件下测定游离HPT的荧光强度，以校正内滤效应。高自旋铁配体，如水和氟，比低自旋氰化高铁血红蛋白的荧光少（更多的是脱氧状态）。正如由于酸碱转变形成羟基高铁血红蛋白所预期的那样，水合高铁血红蛋白样品在pH 7以上显示出向氧合血红蛋白构象的转变。像苯扎贝特这样不与DPG结合在同一部位的效应物，显示出类似脱氧特征的增加。
