Bioorganic Chemistry Laboratory, Molecular Pharmacology and Chemistry Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065.
Proc Natl Acad Sci U S A. 2014 Feb 25;111(8):2891-6. doi: 10.1073/pnas.1400556111. Epub 2014 Feb 10.
Due to the emerging importance of the bromodomain binding region in the study of epigenetic effectors and the vast implications for a wide variety of human disease, the bromodomain region of human ATPase family AAA+ (ATPases associated with diverse cellular activities) domain-containing protein 2 (ATAD2) was targeted for chemical synthesis. The ATAD2 bromodomain (130 aa) was divided into five strategic fragments to be assembled using native chemical ligation with a focus on maximal convergency and efficiency. The fragments were assembled with one cysteine and three thioleucine ligations, unveiling the native alanine and leucine amino acids at the ligation points following metal-free dethiylation. Synthetic highlights of the study are a photolabile dimethoxynitrobenzyl-protected glutamic acid side chain used to impede hydrolysis of the C-terminal Glu-thioester, a thiazolidine-protected thioleucine, and an efficient assembly of three fragments in a single reaction vessel with dual-mode kinetic-standard chemical ligation. With a focus on material throughput and convergency, the five peptide fragments were assembled into the native ATAD2 bromodomain region with a total of three HPLC events in 8% overall yield from the fragments.
由于溴结构域结合区在表观遗传效应物研究中的重要性不断凸显，以及其对多种人类疾病的广泛影响，人们针对人类 ATP 酶家族 AAA+（与多种细胞活动相关的 ATP 酶）域包含蛋白 2（ATAD2）的溴结构域区域进行了化学合成。将 ATAD2 溴结构域（130 个氨基酸）分成五个战略片段，使用天然化学连接进行组装，重点是最大化收敛性和效率。这些片段通过一个半胱氨酸和三个硫代亮氨酸连接进行组装，在无金属脱硫后，在连接点揭示了天然的丙氨酸和亮氨酸氨基酸。该研究的合成亮点包括使用光不稳定的二甲氧基硝基苄基保护的谷氨酸侧链来阻止 C 末端 Glu-硫酯的水解，使用噻唑烷保护的硫代亮氨酸，以及在单个反应容器中以双模式动力学标准化学连接高效组装三个片段。重点关注材料通量和收敛性，将五个肽片段组装成天然的 ATAD2 溴结构域区域，总共进行了三次 HPLC 实验，从片段开始的总收率为 8%。
