Tropomodulin 3 binds to actin monomers
Tropomodulin 3 binds to actin monomers
复制标题
DOI：
10.1074/jbc.m606315200
复制
发表时间：
2006-11-24
影响因子：
4.8
通讯作者：
Fowler, Velia M.
中科院分区：
文献类型：
--
作者：
Fischer, Robert S.;Yarmola, Elena G.;Fowler, Velia M.
微丝覆盖蛋白对肌动蛋白细胞骨架的调节对细胞的各种动态功能至关重要。这些蛋白质结合细丝和单体的能力通常是它们细胞功能的核心。Tmod3是一种普遍存在的尖端封端蛋白，是细胞迁移的负性调节因子，但其细胞功能的机制尚不清楚。对Tmod3对肌动蛋白聚合动力学和稳态单体水平的影响的分析表明，Tmod3不同于以前描述的原模蛋白，它以与其对末端封端的亲和力相似的亲和力隔离肌动蛋白单体。此外，在高速上清胞浆抽提液中发现Tmod3与肌动蛋白结合，这表明Tmod3可以在其他胞浆单体结合蛋白的背景下与单体结合。Tmod3-肌动蛋白复合体可以有效地与1-ethyl-3-(dimethylaminopropyl)carbodiimide/N-hydroxylsulfosuccinimide以1：1的复合体进行交联。随后的胰酶消化和液相色谱/串联质谱仪发现了肌动蛋白上的两个结合界面，一个不同于其他肌动蛋白单体结合蛋白，以及Tmod3中两个潜在的结合部位，它们与先前描述的参与末端封顶的富含亮氨酸的重复结构无关。这些数据表明，Tmod3亚型可能以不同的方式调节细胞内肌动蛋白的动态，而不同于先前描述的原调节蛋白亚型。
Regulation of the actin cytoskeleton by filament capping proteins is critical to myriad dynamic cellular functions. The ability of these proteins to bind both filaments as well as monomers is often central to their cellular functions. The ubiquitous pointed end capping protein Tmod3 ( tropomodulin 3) acts as a negative regulator of cell migration, yet mechanisms behind its cellular functions are not understood. Analysis of Tmod3 effects on kinetics of actin polymerization and steady state monomer levels revealed that Tmod3, unlike previously characterized tropomodulins, sequesters actin monomers with an affinity similar to its affinity for capping pointed ends. Furthermore, Tmod3 is found bound to actin in high speed supernatant cytosolic extracts, suggesting that Tmod3 can bind to monomers in the context of other cytosolic monomer binding proteins. The Tmod3-actin complex can be efficiently cross-linked with 1-ethyl-3-(dimethylaminopropyl)carbodiimide/N-hydroxylsulfosuccinimide in a 1: 1 complex. Subsequent tryptic digestion and liquid chromatography/tandem mass spectrometry revealed two binding interfaces on actin, one distinct from other actin monomer binding proteins, and two potential binding sites in Tmod3, which are independent of the previously characterized leucine-rich repeat structure involved in pointed end capping. These data suggest that the Tmod3 isoform may regulate actin dynamics differently in cells than the previously described tropomodulin isoforms.