Kieffer N, Guichard J, Farcet J P, Vainchenker W, Breton-Gorius J
Eur J Biochem. 1987 Apr 1;164(1):189-95. doi: 10.1111/j.1432-1033.1987.tb11010.x.
We studied de novo protein biosynthesis in platelets of normal adult donors and in newly formed platelets isolated from splenectomized patients with idiopathic thrombocytopenic purpura (ITP). After metabolic labelling of platelet proteins, performed with different radiolabelled amino acids or carbohydrates, a tenfold increase in incorporation of radioactivity into trichloroacetic-acid-precipitable material was obtained with ITP platelets compared to control platelets. Electron microscopic studies of ITP platelets revealed the presence of rough endoplasmic reticulum and polyribosomes, providing morphological evidence for protein synthesis. SDS-PAGE of radiolabelled ITP platelet proteins followed by autoradiography showed that [35S]methionine and [3H]leucine were incorporated into almost all Coomassie-blue-stained proteins whereas [3H]carbohydrates only labelled a few bands. Using crossed-immunoelectrophoresis we identified some of the labelled platelet compounds and demonstrated that major membrane glycoproteins (GPIb, IIb, IIIa) and alpha-granule proteins, such as fibrinogen, thrombospondin, albumin and von Willebrand factor, were synthesized in newly formed circulating platelets.
我们研究了正常成年供血者血小板以及从患有特发性血小板减少性紫癜(ITP)的脾切除患者中分离出的新生血小板中的从头蛋白质生物合成。在用不同的放射性标记氨基酸或碳水化合物对血小板蛋白质进行代谢标记后,与对照血小板相比,ITP血小板中三氯乙酸可沉淀物质的放射性掺入量增加了十倍。对ITP血小板的电子显微镜研究显示存在粗面内质网和多核糖体,为蛋白质合成提供了形态学证据。对放射性标记的ITP血小板蛋白质进行SDS-PAGE并随后进行放射自显影显示,[35S]甲硫氨酸和[3H]亮氨酸几乎掺入了所有考马斯亮蓝染色的蛋白质中,而[3H]碳水化合物仅标记了几条带。使用交叉免疫电泳,我们鉴定了一些标记的血小板化合物,并证明主要的膜糖蛋白(GPIb、IIb、IIIa)和α-颗粒蛋白,如纤维蛋白原、血小板反应蛋白、白蛋白和血管性血友病因子,是在新形成的循环血小板中合成的。