The Cdc42 binding and scaffolding activities of the fission yeast adaptor protein Scd2
The Cdc42 binding and scaffolding activities of the fission yeast adaptor protein Scd2
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DOI：
10.1074/jbc.m209714200
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发表时间：
2003-01-10
影响因子：
4.8
通讯作者：
Yokoyama, S
中科院分区：
文献类型：
--
作者：
Endo, M;Shirouzu, M;Yokoyama, S
小GTP结合蛋白Cdc 42、鸟嘌呤核苷酸交换因子Scd 1、p21激活的激酶Shk 1和衔接蛋白Scd 2参与了裂殖酵母中Cdc 42依赖的信号级联。在本研究中，我们分析了Cdc 42的结合和Scd 2的支架活动的共沉淀法。我们发现，两个SH 3-含有区域，氨基酸残基1-87（CB 1（Cdc 42-binding region 1））和110266（CB 2），的Scd 2可以结合到GTP结合形式的Cdc 42。CB 2是隐蔽的，因为CB 2中的SH 3结构域（SH 3（C））和PX结构域之间的分子内结合，并且仅当Scd 2 PB 1结构域结合到Scdl的含PC基序的区域（残基760-872）时才结合到Cdc 42。这种CB2.Cdc42的结合可以稳定Scd 2的开放构型，使SH 3（C）结构域能够与Shk 1的聚脯氨酸基序结合。我们还发现，GTP结合形式的Cdc 42结合到CRIB基序的Shk 1更强烈比Scd 2。因此，Scd 2作为支架形成蛋白质复合物，并且GTP结合的Cdc 42可以通过Scd 2有效地从上游激活剂Scd 1转移到下游效应物Shk 1。
The small GTP-binding protein Cdc42, the guanine nucleotide exchange factor Scd1, the p21-activated kinase Shk1, and the adaptor protein Scd2 are involved in the Cdc42-dependent signaling cascade in fission yeast. In the present study, we analyzed the Cdc42 binding and scaffolding activities of Scd2 by co-precipitation assays. We found that two SH3-containing regions, amino acid residues 1-87 (CB1 (Cdc42-binding region 1)) and 110266 (CB2), of Scd2 can bind to the GTP-bound form of Cdc42. CB2 is cryptic because of the intramolecular binding between the SH3 domain in CB2 (SH3(C)) and the PX domain and binds to Cdc42 only when the Scd2 PB1 domain binds to the PC motif-containing region (residues 760-872) of Scdl. This CB2.Cdc42 association, which would stabilize the open configuration of Scd2, enables the SH3(C) domain to bind to the polyproline motif of Shk1. We also found that the GTP-bound form of Cdc42 binds to the CRIB motif of Shk1 more strongly than to Scd2. Thus, Scd2 functions as a scaffold to form a protein complex, and the GTP-bound Cdc42 might be transferred effectively from the upstream activator Scdl to the downstream effector Shk1 via Scd2.