Institute of Molecular Biology, University of Oregon, Eugene, Oregon, United States of America.
PLoS One. 2012;7(4):e36014. doi: 10.1371/journal.pone.0036014. Epub 2012 Apr 24.
Membrane Associated Guanylate Kinases (MAGUKs) contain a protein interaction domain (GK(dom)) derived from the enzyme Guanylate Kinase (GK(enz)). Here we show that GK(dom) from the MAGUK Discs large (Dlg) is a phosphoprotein recognition domain, specifically recognizing the phosphorylated form of the mitotic spindle orientation protein Partner of Inscuteable (Pins). We determined the structure of the Dlg-Pins complex to understand the dramatic transition from nucleotide kinase to phosphoprotein recognition domain. The structure reveals that the region of the GK(dom) that once served as the GMP binding domain (GBD) has been co-opted for protein interaction. Pins makes significantly more contact with the GBD than does GMP, but primarily with residues that are conserved between enzyme and domain revealing the versatility of the GBD as a platform for nucleotide and protein interactions. Mutational analysis reveals that the GBD is also used to bind the GK ligand MAP1a, suggesting that this is a common mode of MAGUK complex assembly. The GK(enz) undergoes a dramatic closing reaction upon GMP binding but the protein-bound GK(dom) remains in the 'open' conformation indicating that the dramatic conformational change has been lost in the conversion from nucleotide kinase to phosphoprotein recognition domain.
膜相关鸟苷酸激酶 (MAGUKs) 包含一个来源于鸟苷酸激酶 (GK(enz)) 的蛋白相互作用结构域 (GK(dom))。在这里，我们发现 MAGUKs 中的 Discs large (Dlg) 的 GK(dom) 是一个磷酸化蛋白识别结构域，可特异性识别有丝分裂纺锤体定向蛋白 Partner of Inscuteable (Pins) 的磷酸化形式。我们解析了 Dlg-Pins 复合物的结构，以了解这一从核苷酸激酶到磷酸化蛋白识别结构域的显著转变。结构揭示了 GK(dom) 中曾经作为 GMP 结合域 (GBD) 的区域被用于蛋白相互作用。Pins 与 GBD 的结合比 GMP 更紧密，但主要与酶和结构域之间保守的残基结合，揭示了 GBD 作为核苷酸和蛋白相互作用平台的多功能性。突变分析揭示 GBD 也用于结合 GK 配体 MAP1a，这表明这是 MAGUK 复合物组装的一种常见模式。当 GMP 结合时，GK(enz) 会发生剧烈的关闭反应，但与蛋白结合的 GK(dom) 仍保持“开放”构象，表明在从核苷酸激酶到磷酸化蛋白识别结构域的转变中，这一剧烈的构象变化已经丧失。
