CNRS and Ecole normale supérieure Paris-Saclay, Laboratoire de Biologie et Pharmacologie Appliquée, 61 Avenue du Président Wilson, 94235, Cachan CEDEX, France.
Institut Pasteur and CNRS UMR 3525, Biologie des Bactéries Intracellulaires, 25-28 Rue du Dr Roux, 75015, Paris, France.
Nat Commun. 2019 Mar 8;10(1):1142. doi: 10.1038/s41467-019-09023-1.
FIC proteins regulate molecular processes from bacteria to humans by catalyzing post-translational modifications (PTM), the most frequent being the addition of AMP or AMPylation. In many AMPylating FIC proteins, a structurally conserved glutamate represses AMPylation and, in mammalian FICD, also supports deAMPylation of BiP/GRP78, a key chaperone of the unfolded protein response. Currently, a direct signal regulating these FIC proteins has not been identified. Here, we use X-ray crystallography and in vitro PTM assays to address this question. We discover that Enterococcus faecalis FIC (EfFIC) catalyzes both AMPylation and deAMPylation and that the glutamate implements a multi-position metal switch whereby Mg and Ca control AMPylation and deAMPylation differentially without a conformational change. Remarkably, Ca concentration also tunes deAMPylation of BiP by human FICD. Our results suggest that the conserved glutamate is a signature of AMPylation/deAMPylation FIC bifunctionality and identify metal ions as diffusible signals that regulate such FIC proteins directly.
FIC 蛋白通过催化翻译后修饰（PTM）来调节从细菌到人类的分子过程，最常见的是添加 AMP 或 AMP 化。在许多 AMP 化的 FIC 蛋白中，结构保守的谷氨酸抑制 AMP 化，并且在哺乳动物 FICD 中，也支持 BiP/GRP78 的去 AMP 化，BiP/GRP78 是未折叠蛋白反应的关键伴侣蛋白。目前，尚未鉴定出直接调节这些 FIC 蛋白的信号。在这里，我们使用 X 射线晶体学和体外 PTM 测定来解决这个问题。我们发现粪肠球菌 FIC（EfFIC）催化 AMP 化和去 AMP 化，谷氨酸执行多位置金属开关，其中 Mg 和 Ca 不同地控制 AMP 化和去 AMP 化，而没有构象变化。值得注意的是，Ca 浓度也调节人 FICD 对 BiP 的去 AMP 化。我们的结果表明，保守的谷氨酸是 AMP 化/去 AMP 化 FIC 双功能的特征，并确定金属离子是直接调节此类 FIC 蛋白的可扩散信号。
