Mutational analysis on the function of the SWAP-70 PH domain
Mutational analysis on the function of the SWAP-70 PH domain
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DOI：
10.1007/s11010-006-9236-1
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发表时间：
2006-12-01
影响因子：
4.3
通讯作者：
Fukui, Yasuhisa
中科院分区：
文献类型：
--
作者：
Wakamatsu, Isamu;Ihara, Sayoko;Fukui, Yasuhisa
SWAP-70是一种三磷酸磷脂酰肌醇(PtdIns(3，4，5)P-3)结合蛋白，可能与激活的RAC协同参与膜褶皱。在PH域引入了各种点突变。丙氨酸取代了第一环中带正电荷的氨基酸，取消了PH结构域与PtdIns(3，4，5)P-3的结合活性。PtdIns(3，4，5)P-3结合活性是SWAP-70转位到膜上所必需的，过表达的蛋白增强膜褶皱，或者在膜褶皱中缺乏羧基末端区域的突变体的显性-负效应。当Rac过表达时，上述突变体被转移到膜上，并对膜褶皱表现出显性-负性效应，而不具有PtdIns(3，4，5)P-3结合活性。这些结果表明，当SWAP-70和RAC有效地相互作用时，PtdIns(3，4，5)P3结合活性对于这些事件是必不可少的。这些结果表明，SWAP-70与PtdIns(3，4，5)P-3的结合可能有助于SWAP-70在活化的RAC上的募集。
SWAP-70 is a phosphatidylinositol trisphosphate (PtdIns(3,4,5)P-3)-binding protein, which is suggested to be involved in membrane ruffling, cooperating with activated Rac. Various point mutations were introduced in the PH domain. Substitutions of alanines for the positively charged amino acids within the first loop abolished the binding activity of the PH domains to PtdIns(3,4,5)P-3. The PtdIns(3,4,5)P-3 binding activity was required for translocation of SWAP-70 to the membrane, enhancement of membrane ruffling by the overexpressed protein, or the dominant-negative effect of a mutant lacking the carboxyl terminal region in membrane ruffling. When Rac was overexpressed, the above mutants were translocated to the membrane and exhibited a dominant-negative effect on membrane ruffling without PtdIns(3,4,5)P-3-binding activity. These results suggest that the PtdIns(3,4,5) P3-binding activity is dispensable for these events when SWAP-70 and Rac interacts efficiently. These results implicate that binding of SWAP-70 to PtdIns(3,4,5)P-3 may facilitate the recruitment of SWAP-70 to activated Rac.