Kinetics of Src Homology 3 Domain Association with the Proline-rich Domain of Dynamins
Kinetics of Src Homology 3 Domain Association with the Proline-rich Domain of Dynamins
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Src 同源 3 结构域与富含脯氨酸的 Dynamins 结构域关联的动力学
DOI：
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发表时间：
2005
影响因子：
4.8
通讯作者：
H. Palfrey
中科院分区：
文献类型：
--
作者：
E. Solomaha;Frances L. Szeto;Mohammed A. Yousef;H. Palfrey
动力蛋白的功能部分是通过其富含脯氨酸结构域（PRD）与几种假定结合蛋白的Src同源3 （SH3）结构域的关联而介导的。为了评估这一过程的特异性和动力学，我们对分离的动力蛋白-1和-2的PRDs与几个纯化的SH3结构域之间的相互作用进行了表面等离子体共振研究。谷胱甘肽s -转移酶连接的SH3结构域与dynamin-1和-2具有高亲和力（KD ~ 10 nm ~ 1 μm）。最简单的相互作用似乎发生在amphiphysin-SH3结构域；这与先前研究预测的dynamin-1 PRD C端单一高亲和力位点（KD ~ 10 nm）结合。与动力蛋白-2 PRD的结合也是单相的，但亲和力略低（KD ~ 25 nm）。Endophilin-SH3与dynamin-1和-2 PRD的结合是双相的，在PRD的N端有一个高亲和力位点（KD ~ 14 nm），在dynamin-1的C端有一个低亲和力位点（KD ~ 60 nm）。dynamin-2 PRD的n端位点对内啡肽sh3的亲和力降低了10倍。带amphiphysin-SH3结构域的dynamin-1 PRD的预加载部分阻断了内啡肽- sh3结构域的结合，表明C端结合位点之间存在重叠，但内啡肽仍然能够与高亲和力的n端位点相互作用。这表明不止一个SH3结构域可以同时与PRD结合，并表明体内不同的含SH3蛋白之间可能发生竞争，争夺有限数量的PXXP基序。当动力蛋白-1 PRD被Cdk5磷酸化时，内啡肽- sh3与高亲和力位点的结合被破坏，表明该位点与磷酸化位点重叠，但amphiphysin-SH3的结合不受影响。其他SH3结构域也表现出类似的复杂结合特征，dynamin-1和-2的prd之间存在显著差异。例如，来自c-Src、Grb2和intersectin的SH3结构域仅与dynamin-2 PRD的c端部分结合，而与dynamin-1 PRD的N端和c端部分结合。因此，含有SH3结构域的蛋白与动力蛋白-1和-2的不同结合可能有助于这些同工异构体发挥不同的功能。
Dynamin function is mediated in part through association of its proline-rich domain (PRD) with the Src homology 3 (SH3) domains of several putative binding proteins. To assess the specificity and kinetics of this process, we undertook surface plasmon resonance studies of the interaction between isolated PRDs of dynamin-1 and -2 and several purified SH3 domains. Glutathione S-transferase-linked SH3 domains bound with high affinity (KD ∼10 nm to 1 μm) to both dynamin-1 and -2. The simplest interaction appeared to take place with the amphiphysin-SH3 domain; this bound to a single high affinity site (KD ∼10 nm) in the C terminus of dynamin-1 PRD, as predicted by previous studies. Binding to the dynamin-2 PRD was also monophasic but with a slightly lower affinity (KD ∼25 nm). Endophilin-SH3 binding to both dynamin-1 and -2 PRDs was biphasic, with one high affinity site (KD ∼14 nm) in the N terminus of the PRD and another lower affinity site (KD ∼60 nm) in the C terminus of dynamin-1. The N-terminal site in dynamin-2 PRD had a 10-fold lower affinity for endophilin-SH3. Preloading of dynamin-1 PRD with the amphiphysin-SH3 domain partially occluded binding of the endophilin-SH3 domain, indicating overlap between the binding sites in the C terminus, but endophilin was still able to interact with the high affinity N-terminal site. This shows that more than one SH3 domain can simultaneously bind to the PRD and suggests that competition probably occurs in vivo between different SH3-containing proteins for the limited number of PXXP motifs. Endophilin-SH3 binding to the high affinity site was disrupted when dynamin-1 PRD was phosphorylated with Cdk5, indicating that this site overlaps the phosphorylation sites, but amphiphysin-SH3 binding was unaffected. Other SH3 domains showed similarly complex binding characteristics, and substantial differences were noted between the PRDs from dynamin-1 and -2. For example, SH3 domains from c-Src, Grb2, and intersectin bound only to the C-terminal half of dynamin-2 PRD but to both the N- and C-terminal portions of dynamin-1 PRD. Thus, differential binding of SH3 domain-containing proteins to dynamin-1 and -2 may contribute to the distinct functions performed by these isoforms.
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影响因子：
4
作者：
B. Mayer
通讯作者：
B. Mayer
影响因子：
56.9
作者：
FENG, SB;CHEN, JK;SCHREIBER, SL
通讯作者：
SCHREIBER, SL
DOI：
10.1073/pnas.94.16.8569
发表时间：
1997-08-05
影响因子：
11.1
作者：
Ringstad, N;Nemoto, Y;DeCamilli, P
通讯作者：
DeCamilli, P
影响因子：
2.9
作者：
Dutta, K;Shi, HH;Ghose, R
通讯作者：
Ghose, R
影响因子：
56.9
作者：
Slepnev, VI;Ochoa, GC;De Camilli, P
通讯作者：
De Camilli, P