Bertolucci Craig M, Guibao Cristina D, Zheng Jie J
Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
Biochemistry. 2008 Jan 15;47(2):548-54. doi: 10.1021/bi702103n. Epub 2007 Dec 20.
Cell migration is a dynamic process that requires the coordinated formation and disassembly of focal adhesions (FAs). Several proteins such as paxillin, focal adhesion kinase (FAK), and G protein-coupled receptor kinase-interacting protein 1 (GIT1) are known to play a regulatory role in FA disassembly and turnover. However, the mechanisms by which this occurs remain to be elucidated. Paxillin has been shown to bind the C-terminal domain of FAK in FAs, and an increasing number of studies have linked paxillin association with GIT1 during focal adhesion disassembly. It has been reported recently that phosphorylation of serine 273 in the LD4 motif of paxillin leads to an increased association with Git1 and focal adhesion turnover. In the present study, we examined the effects of phosphorylation of the LD4 peptide on its binding affinity to the C-terminal domain of FAK. We show that phosphorylation of LD4 results in a reduction of binding affinity to FAK. This reduction in binding affinity is not due to the introduction of electrostatic repulsion or steric effects but rather by a destabilization of the helical propensity of the LD4 motif. These results further our understanding of the focal adhesion turnover mechanism as well as identify a novel process by which phosphorylation can modulate intracellular signaling.
细胞迁移是一个动态过程，需要粘着斑（FAs）的协调形成和解聚。已知几种蛋白质，如桩蛋白、粘着斑激酶（FAK）和G蛋白偶联受体激酶相互作用蛋白1（GIT1）在粘着斑的解聚和周转中发挥调节作用。然而，其发生机制仍有待阐明。桩蛋白已被证明在粘着斑中与FAK的C末端结构域结合，并且越来越多的研究将粘着斑解聚过程中桩蛋白与GIT1的结合联系起来。最近有报道称，桩蛋白LD4基序中丝氨酸273的磷酸化导致与Git1的结合增加以及粘着斑周转。在本研究中，我们研究了LD4肽磷酸化对其与FAK C末端结构域结合亲和力的影响。我们发现LD4的磷酸化导致与FAK的结合亲和力降低。这种结合亲和力的降低不是由于引入了静电排斥或空间效应，而是由于LD4基序螺旋倾向的不稳定。这些结果进一步加深了我们对粘着斑周转机制的理解，并确定了一种磷酸化可调节细胞内信号传导的新过程。
