Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, UMR 7099 CNRS, Université Paris Diderot, Sorbonne Paris Cité, PSL Research University, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, 75005, Paris, France.
Institut Jacques Monod UMR 7592, CNRS, Université Paris Diderot, Sorbonne Paris Cité, F-75205, Paris, France.
Sci Rep. 2017 Sep 11;7(1):11139. doi: 10.1038/s41598-017-11203-2.
Suppressor of Fused (SUFU) is a highly conserved protein that acts as a negative regulator of the Hedgehog (HH) signalling pathway, a major determinant of cell differentiation and proliferation. Therefore, SUFU deletion in mammals has devastating effects on embryo development. SUFU is part of a multi-protein cytoplasmic signal-transducing complex. Its partners include the Gli family of transcription factors that function either as repressors, or as transcription activators according to the HH activation state. The crystal structure of SUFU revealed a two-domain arrangement, which undergoes a closing movement upon binding a peptide from Gli1. There remains however, much to be discovered about SUFU's behaviour. To this end, we expressed recombinant, full-length SUFU from Drosophila, Zebrafish and Human. Guided by a sequence analysis that revealed a conserved potential metal binding site, we discovered that SUFU binds zinc. This binding was found to occur with a nanomolar affinity to SUFU from all three species. Mutation of one histidine from the conserved motif induces a moderate decrease in affinity for zinc, while circular dichroism indicates that the mutant remains structured. Our results reveal new metal binding affinity characteristics about SUFU that could be of importance for its regulatory function in HH.
抑制融合蛋白（SUFU）是一种高度保守的蛋白质，作为 Hedgehog（HH）信号通路的负调节剂，在细胞分化和增殖中起着重要作用。因此，哺乳动物中 SUFU 的缺失对胚胎发育有毁灭性的影响。SUFU 是一种多蛋白细胞质信号转导复合物的一部分。它的伴侣包括 Gli 家族转录因子，根据 HH 的激活状态，这些转录因子作为抑制剂或转录激活剂发挥作用。SUFU 的晶体结构揭示了一个双结构域排列，在结合来自 Gli1 的肽后会发生闭合运动。然而，关于 SUFU 的行为仍有很多需要发现。为此，我们从果蝇、斑马鱼和人类中表达了重组全长 SUFU。通过序列分析揭示了一个保守的潜在金属结合位点的指导，我们发现 SUFU 结合锌。从所有三个物种中发现 SUFU 与锌的结合具有纳摩尔亲和力。从保守基序中突变一个组氨酸会导致锌结合亲和力适度降低，而圆二色性表明突变体仍然具有结构。我们的结果揭示了 SUFU 的新的金属结合亲和力特征，这可能对其在 HH 中的调节功能很重要。
