Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabe-dori, Mizuho-ku, Nagoya 467-8603, Japan.
Biochem Biophys Res Commun. 2010 Mar 12;393(3):487-91. doi: 10.1016/j.bbrc.2010.02.030. Epub 2010 Feb 10.
Osteopontin (OPN) is an integrin-binding protein found in a variety of tissues and physiological fluids and is involved in divergent biological processes such as migration, adhesion and signaling in integrin-independent as well as dependent manners. The adhesive activity of this protein is modulated upon cleavage by thrombin at the central part of the molecule, in the vicinity of the integrin-binding sequences. Although detailed structural characterization is crucial for further understanding of the regulatory mechanisms of the OPN functions, its intrinsically disordered property hampers in-depth conformational analyses. Here we report an NMR study of mouse OPN and its N-terminal thrombin-cleavage product to characterize intramolecular interaction of this molecule. Paramagnetic relaxation enhancement experiment revealed that OPN exhibits a long-range intramolecular interaction between the N- and C-terminal regions. Furthermore, our NMR data showed that anti-OPN antibody OPN1.2, whose reactivity is impaired by deletion or amino acid substitutions of the arginine-aspartate-glycine integrin-binding motif, binds the N-terminal side of the integrin-binding motifs suggesting the existence of intramolecular interaction. These data suggest that functional interactions of OPN with integrins and the other binding partners can be modulated by the intramolecular interactions.
骨桥蛋白（OPN）是一种整合素结合蛋白，存在于多种组织和生理液中，参与迁移、黏附和信号转导等多种生物学过程，整合素独立和依赖的方式。该蛋白的黏附活性在分子中部被凝血酶切割时发生调节，靠近整合素结合序列。尽管详细的结构特征对于进一步理解 OPN 功能的调控机制至关重要，但该蛋白的固有无序特性阻碍了深入的构象分析。本研究通过核磁共振（NMR）研究了小鼠 OPN 及其 N 端凝血酶切割产物，以表征该分子的分子内相互作用。顺磁松弛增强实验表明，OPN 分子的 N 端和 C 端区域之间存在长程分子内相互作用。此外，我们的 NMR 数据表明，抗 OPN 抗体 OPN1.2 的反应性受到精氨酸-天冬氨酸-甘氨酸整合素结合基序缺失或氨基酸取代的损害，结合整合素结合基序的 N 端侧，表明存在分子内相互作用。这些数据表明，OPN 与整合素和其他结合伴侣的功能相互作用可以通过分子内相互作用来调节。
