Subcellular localization of a thromboxane A2/prostaglandin H2 receptor antagonist binding site in human platelets.
Subcellular localization of a thromboxane A2/prostaglandin H2 receptor antagonist binding site in human platelets.
复制标题
人血小板中血栓素 A2/前列腺素 H2 受体拮抗剂结合位点的亚细胞定位。
DOI：
10.1016/0006-2952(88)90138-4
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发表时间：
1988
影响因子：
5.8
通讯作者：
Halushka,PV
中科院分区：
文献类型：
--
作者：
SaussyJr,DL;Mais,DE;Baron,DA;Pepkowitz,SH;Halushka,PV
测定了竞争性血栓素A_2/前列腺素H_2（TXA_2/PGH_2）拮抗剂9，11-二甲基亚甲基-11，12-亚甲基-16-（3-碘-4-羟基-苯基）-13，14-二氢-13-氮杂-15 αβ-ω-四去甲TXA_2（[125 I]-PTA-OH）结合位点的亚细胞定位。通过甘油裂解或氮气空化制备血小板的亚细胞组分，并通过使用质膜、内质网（致密管状系统）、线粒体、颗粒和胞质组分特异性的酶标记物进行表征。通过Scatchard平衡结合数据分析确定亚细胞组分中结合位点的Kd和密度。在裂解物中测定的[125 I]-PTA-OH的Kd和Bmax分别为49 ± 11 nM和4.1 ± 1.7 pmol/mg蛋白（N = 6）。在测定的任何组分中，Kd值均无显著差异。结合位点共富集（4.5 ± 0.66倍），其中质膜酶标记物共富集（3.7 ± 0.5倍），致密小管酶标记物共富集（2.4 ± 0.4倍）。结合位点没有与细胞质成分、线粒体或颗粒的标记物共同富集。TXA 2/PGH 2模拟物U46619与[125 I]-PTA-OH竞争结合位点的能力也被确定为各种亚细胞级分。U46619在裂解液中的IC 50为5.4 ± 1.2μM，在亚细胞组分中无显著差异。这些数据表明，结合位点是上述TXA_2/PGH_2受体。这些数据与推测的TXA_2/PGH_2受体定位于质膜和/或致密小管系统的观点一致。
The subcellular localization of a binding site for the competitive thromboxane A2/prosta-glandin H2(TXA2/PGH2) antagonist, 9,11-dimethylmethano-11,12-methano-16-(3-iodo-4-hydroxy-phenyl)-13,14-dihydro-13-aza-15αβ-ω-tetranor TXA2([125I]-PTA-OH), was determined. Subcellular fractions of platelets were prepared by glycerol lysis or nitrogen cavitation, and were characterized by the use of enzymatic markers specifie for plasma membranes, endoplasmic reticulum (dense tubular system), mitochondria, granules, and cytosolic constituents. TheKdand density of binding sites in the subcellular fractions were determined by Scatchard analysis of equilibrium binding data. TheKdandBmaxfor [125I]-PTA-OH determined in the lysates were 49 ± 11 nM and 4.1 ± 1.7 pmol/mg protein respectively (N = 6). TheKdvalues were not significantly different in any of the fractions assayed. The binding sites were coenriched (4.5 ± 0.66 fold) with the enzymatic markers for plasma membranes (3.7 ± 0.5 fold) and dense tubular system (2.4 ± 0.4 fold). The binding sites were not coenriched with markers for cytoplasmic constituents, mitochondria, or granules. The ability of the TXA2/PGH2mimetic U46619 to compete with [125I]-PTA-OH for the binding site was also determined for the various subcellular fractions. TheIC50for U46619 was 5.4 ± 1.2μM in the lysate, and was not significantly different in the subcellular fractions. These data suggest that the binding site is the TXA2/PGH2receptor described previously. These data are consistent with the notion that the putative TXA2/PGH2receptor is localized in the plasma membranes and/or the dense tubular system.