Different Roles of N- and C-Termini in the Functional Activity of FGF21
Different Roles of N- and C-Termini in the Functional Activity of FGF21
复制标题
DOI：
10.1002/jcp.21675
复制
发表时间：
2009-05-01
影响因子：
5.6
通讯作者：
Kharitonenkov, Alexei
中科院分区：
文献类型：
--
作者：
Micanovic, Radmila;Raches, David W.;Kharitonenkov, Alexei
成纤维细胞生长因子21是与FGF19、FGF23一起属于内分泌FGFs亚家族的成员。它正在成为一种新的调节剂，对包括血糖和血脂控制在内的各种代谢参数都有有益的影响。FGF21的活性依赖于膜蛋白βKlotho，该蛋白与不同的FGF21受体形成物理络合物，从而赋予它们结合FGF21并激活下游信号通路的能力。FGF21和其他FGFs一样，折叠成一个P-三叶形的核心区，N-末端和C-末端无序。为了研究它们在FGF21活性中的作用，我们构建了一系列缺失突变体，并检测了它们(1)结合β-Klotho的能力，(2)通过MAPK磷酸化分析它们与β-Klotho结合的能力；(2)通过MAPK磷酸化分析它们抑制3T3-Li/beta Klotho成纤维细胞的凋亡；(3)刺激3T3-Li脂肪细胞GLUTI基因的表达和葡萄糖摄取。与β-Klotho的结合研究表明，与辅助受体的相互作用涉及C-末端，因为从羧基末端逐渐去除氨基酸降低了与β-Klotho的亲和力。相比之下，从N-末端移除多达17个氨基酸对与β-Klotho的相互作用没有影响。末端缺失对功能的影响更大，因为氨基端的6个氨基酸和羧端的4个氨基酸的缺失对活性都有显著影响(10倍)。在没有检测到活性的极端末端截断中，AN 17作为竞争性拮抗剂，而Delta C20不起作用。我们的结构/功能研究表明，C-末端对β-Klotho相互作用很重要，而N-末端可能直接与成纤维细胞生长因子受体相互作用。
Fibroblast growth factor 21 is a member of endocrine FGFs subfamily, along with FGF19 and FGF23. It is emerging as a novel regulator with beneficial effects on a variety of metabolic parameters, including glucose and lipid control. FGF21 activity depends on membrane protein beta Klotho that physically complexes with various FGF receptors, thus conferring them the ability to bind FGF21 and activate downstream signaling pathways. FGF21, like other FGFs, folds to a P-trefoil-like core region, with disordered N- and C-termini. In order to investigate their role in the activity of FGF21, we have constructed a series of deletion mutants and tested them for their ability to (1) bind beta Klotho, analyzed by surface plasmon resonance spectroscopy (2) signal through MAPK phosphorylation and inhibit apoptosis in 3T3-LI/beta Klotho fibroblasts (3) stimulate GLUTI mRNA upregulation and glucose uptake in 3T3-LI adipocytes. Binding studies with beta Klotho revealed that the interaction with the co-receptor involves the C-terminus, as progressive removal of amino acids from the carboxy end decreased affinity for beta Klotho. By contrast, removal of up to 17 amino acids from the N-terminus had no effect on the interaction with beta Klotho. Terminal deletions had greater effect on function, as deletions of six amino acids from the amino-terminus and only four from the carboxy-terminus each significantly impacted activity (10-fold). Of the extreme terminal truncations, with no detectable activity, AN 17 acted as competitive antagonist while Delta C20 did not. Our structure/function studies show that the C-terminus is important for beta Klotho interaction whereas the N-terminus likely interacts directly with FGF receptors.