血小板中肌动蛋白结合蛋白的磷酸化程度与它的肌动蛋白结合能力之间关系及其部分氨基酸顺序

PARTIAL AMINO ACID SEQUENCES OF HUMAN PLATELET ACTIN BINDIN PROTEIN, AND THE RELATIONSHIP BETWEEN ITS PHOSPHORYLA-TION AND ACTIN BINDING ABILITY

我们以前的研究显示人血小板中存在4种磷酸化程度不同的肌动蛋白结合蛋白:ABP0、ABP1、ABP2和ABP3。电泳扫描结果表明:完全磷酸化的ABP3结合肌动蛋白能力最强,约为部分磷酸化的ABP(ABP1和ABP2)的2倍。ABP1和ABP2具有相似结合能力,而非磷酸化的ABP0几乎失去结合肌动蛋白的能力。利用电泳洗脱法我们分离到电泳纯的ABP及其他的钙依赖巯基蛋白酶(CDSP)分解产物,Mr190×10~3、90×10~8两个片段。氨基酸顺序分析结果显示ABP分子的N-端被封闭,190×10~3片段N-端的前1个氨基酸顺序为ATEKDLAE OAP,与人平滑肌细胞的细丝蛋白分子中T110片段相同。 以上结果提示:(1)cAMP调控蛋白激酶(PKA)和蛋白激酶G(PKC)的磷酸化对ABP结合肌动蛋白是必须的;(2)ABP结合肌动蛋白的能力随磷酸化程度增强而增加;(3)血小板和平滑肌细胞的肌动蛋白结合蛋白可能具有同源性。

Our previous study has shown that there are four kinds of act in binding protein(ABP) in the human blood platelet, ABP0, ABP1, ABP2and ABP3. which are phosphorylated to different extents. The scanning results of SDS-PAGE of ABP-aotin complexes show that the binding ability of ABP3 to aotin is strongest, ABPO has no aotin-binding ability. ABP1 and ABP2 bind to actin to similar extent.Using an eleotroeluting technique, we have successfully purified ABP and its proteolytic products, Mr 190 ×103 and 90 ×103 fragments, by calcium-dependent sulfhydryl protease (CDSP). Results of amino acid sequencing show that the N-terminal of ABP is blocked, but the 190 ×103 fragment has an N-terminal sequece of ATEKDLAEDAP, which is the same as the T110 fragment in human smooth muscle filamin.These results together with our previous results suggest that: (1) It is neccessary for binding to actin that ABP is phosphorylated by PKO and PKA. (2) The binding ability of ABP to aotin increases with the extent of its phosph-orylation. (3) Aotin-binding proteins in the human blood platelet and smooth muscle could be homologous.