Lee Wei-Chao, Lin Wen-Ling, Matsui Tsutomu, Chen Eric S-W, Wei Tong-You Wade, Lin Wen-Hsuan, Hu Hao, Zheng Yujun George, Tsai Ming-Daw, Ho Meng-Chiao
Institute of Biological Chemistry, Academia Sinica , Taipei 115, Taiwan.
Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, Stanford University , Menlo Park, California 94025, United States.
Biochemistry. 2015 Dec 29;54(51):7514-23. doi: 10.1021/acs.biochem.5b00995. Epub 2015 Dec 15.
Type I protein arginine methyltransferases (PRMTs) catalyze asymmetric dimethylation of various proteins, and their dysregulations often correlate with tumorigenesis or developmental deficiency. Recent studies have focused on the in vivo substrate identification and the enzyme mechanism with peptide substrates. However, how PRMTs recognize substrates at the protein level remains unknown. PRMT8 is one of the least characterized type I PRMTs, and its crystal structure has not been reported. Here, we report the crystal structure of the PRMT8:SAH complex, identify a new non-histone protein substrate NIFK, and uncover a previously unknown regulatory region specifically required for recognizing NIFK. Instead of the canonical dimeric structure for other type I PRMTs, PRMT8 exists as a tetramer in solution. Using X-ray crystallography in combination with small-angle X-ray scattering experiments, the dimer of dimers architecture in which two PRMT8 dimers are held together mainly by β strand interactions was proposed. Mutation of PRMT8-β15 impedes the methylation of NIFK but still allows methylation of the histone H2A/H2B dimer or a peptide substrate, suggesting a possible structural basis for recognition of protein substrates. Lastly, we observed two PRMT8 dimer orientations resulting in open (without SAH) and closed (with SAH bound) conformations. The comparison between open and closed conformations may provide useful information for PRMT1/8 inhibitor design.
I型蛋白质精氨酸甲基转移酶（PRMTs）催化多种蛋白质的不对称二甲基化，其失调往往与肿瘤发生或发育缺陷相关。最近的研究集中在体内底物鉴定以及肽底物的酶作用机制上。然而，PRMTs如何在蛋白质水平上识别底物仍不清楚。PRMT8是特征最少的I型PRMTs之一，其晶体结构尚未见报道。在此，我们报道了PRMT8:SAH复合物的晶体结构，鉴定出一种新的非组蛋白底物NIFK，并揭示了识别NIFK所特有的一个以前未知的调节区域。与其他I型PRMTs的典型二聚体结构不同，PRMT8在溶液中以四聚体形式存在。结合X射线晶体学和小角X射线散射实验，提出了由两条β链相互作用将两个PRMT8二聚体结合在一起的二聚体-二聚体结构。PRMT8-β15的突变会阻碍NIFK的甲基化，但仍允许组蛋白H2A/H2B二聚体或肽底物的甲基化，这表明了识别蛋白质底物的可能结构基础。最后，我们观察到两种PRMT8二聚体取向，分别导致开放（无SAH）和封闭（结合SAH）构象。开放和封闭构象之间的比较可能为PRMT1/8抑制剂的设计提供有用信息。
