Allosteric Interactions Direct Binding and Phosphorylation of ASF/SF2 by SRPK1
Allosteric Interactions Direct Binding and Phosphorylation of ASF/SF2 by SRPK1
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DOI：
10.1021/bi901107q
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发表时间：
2009-12-08
期刊：
影响因子：
2.9
通讯作者：
Ghosh, Gourisankar
中科院分区：
文献类型：
--
作者：
Huynh, Nhat;Ma, Chen-Ting;Ghosh, Gourisankar
ASF/SF 2是丝氨酸-精氨酸（SR）蛋白家族的成员，具有两个RRM结构域（RRM 1和RRM 2）和一个富含RS二肽的C端结构域。SR蛋白激酶1（SRPK 1）使用半进行性机制磷酸化这些丝氨酸中的约12个。ASF SF 2-SRPK 1复合物的X射线结构揭示了复合物的几个特征，这些特征提出了关于底物如何被激酶磷酸化的有趣问题。RS结构域的在反应的后期阶段注定被磷酸化的部分对接到活性位点远端的激酶沟，而相邻的RRM 2在活性位点附近结合[Ngo，J.C.，等人（2008）Mol. Cell 29，563-576]。在这项研究中，我们研究了RS结构域和RRM 2之间的相互作用，以稳定的协会和ASF/SF 2的磷酸化。尽管在酶-底物复合物中有几个接触，游离RRM 2不能有效地结合SRPK 1，除非对接槽被RS结构域占据。这种结构域串扰增强了RS结构域的进行性磷酸化。RRM-SRPK 1接触残基控制RRM 2中关键β链的折叠。该结构元件的解折叠可迫使RS结构域的N-末端丝氨酸进入活性位点进行顺序磷酸化。因此，ASF/SF 2代表了一类新的底物，其使用独特的一级序列来诱导变构结合、进行性磷酸化和产物释放。
ASF/SF2, a member of the serine-arginine (SR) protein family, has two RRM domains (RRM1 and RRM2) and a C-terminal domain rich in RS dipeptides. SR protein kinase 1 (SRPK1) phosphorylates approximately 12 of these serines using a semiprocessive mechanism. The X-ray structure of the ASF SF2-SRPK1 complex revealed several features of the complex that raised intriguing questions about how the substrate is phosphorylated by the kinase. The part of the RS domain destined to be phosphorylated at later stages of the reaction docks to a kinase groove distal to the active site while the neighboring RRM2 binds near the active site [Ngo, J. C., et al. (2008) Mol. Cell 29, 563-576]. In this study, we investigate the interplay between the RS domain and RRM2 for stable association and phosphorylation of ASF/SF2. Despite several contacts in the enzyme-substrate complex, free RRM2 does not bind efficiently to SRPK1 unless the docking groove is occupied by the RS domain. This domain cross-talk enhances the processive phosphorylation of the RS domain. The RRM-SRPK1 contact residues control the folding of a critical beta-strand in RRM2. Unfolding of this structural element may force the N-terminal serines of the RS domain into the active site for sequential phosphorylation. Thus, ASF/SF2 represents a new class of substrates that use unique primary sequence to induce allosteric binding, processive phosphorylation, and product release.