Dong Cheng, Mao Yunfei, Tempel Wolfram, Qin Su, Li Li, Loppnau Peter, Huang Rong, Min Jinrong
Structural Genomics Consortium, University of Toronto, Toronto, Ontaria M5G 1L7, Canada;
Department of Medicinal Chemistry, Drug Discovery and Development, Virginia Commonwealth University, Richmond, Virginia 23219, USA; The Institute for Structural Biology, Drug Discovery and Development, Virginia Commonwealth University, Richmond, Virginia 23219, USA;
Genes Dev. 2015 Nov 15;29(22):2343-8. doi: 10.1101/gad.270611.115. Epub 2015 Nov 5.
α-N-terminal methylation represents a highly conserved and prevalent post-translational modification, yet its biological function has remained largely speculative. The recent discovery of α-N-terminal methyltransferase 1 (NTMT1) and its physiological substrates propels the elucidation of a general role of α-N-terminal methylation in mediating DNA-binding ability of the modified proteins. The phenotypes, observed from both NTMT1 knockdown in breast cancer cell lines and knockout mouse models, suggest the potential involvement of α-N-terminal methylation in DNA damage response and cancer development. In this study, we report the first crystal structures of human NTMT1 in complex with cofactor S-adenosyl-L-homocysteine (SAH) and six substrate peptides, respectively, and reveal that NTMT1 contains two characteristic structural elements (a β hairpin and an N-terminal extension) that contribute to its substrate specificity. Our complex structures, coupled with mutagenesis, binding, and enzymatic studies, also present the key elements involved in locking the consensus substrate motif XPK (X indicates any residue type other than D/E) into the catalytic pocket for α-N-terminal methylation and explain why NTMT1 prefers an XPK sequence motif. We propose a catalytic mechanism for α-N-terminal methylation. Overall, this study gives us the first glimpse of the molecular mechanism of α-N-terminal methylation and potentially contributes to the advent of therapeutic agents for human diseases associated with deregulated α-N-terminal methylation.
α-N-末端甲基化是一种高度保守且普遍存在的翻译后修饰，但其生物学功能在很大程度上仍属推测。最近发现的α-N-末端甲基转移酶1（NTMT1）及其生理底物推动了对α-N-末端甲基化在介导修饰蛋白的DNA结合能力中普遍作用的阐明。从乳腺癌细胞系中NTMT1敲低以及基因敲除小鼠模型中观察到的表型表明，α-N-末端甲基化可能参与DNA损伤反应和癌症发展。在本研究中，我们分别报道了人NTMT1与辅因子S-腺苷-L-高半胱氨酸（SAH）以及六种底物肽形成复合物的首个晶体结构，并揭示NTMT1包含两个有助于其底物特异性的特征性结构元件（一个β发夹和一个N末端延伸）。我们的复合物结构，结合诱变、结合和酶学研究，还展示了将共有底物基序XPK（X表示除D/E以外的任何残基类型）锁定到催化口袋中进行α-N-末端甲基化所涉及的关键元件，并解释了为什么NTMT1更喜欢XPK序列基序。我们提出了α-N-末端甲基化的催化机制。总体而言，这项研究让我们首次了解了α-N-末端甲基化的分子机制，并可能有助于开发与α-N-末端甲基化失调相关的人类疾病治疗药物。
