Institute of Biochemistry, University of Muenster, Wilhelm-Klemm-Str. 2, D-48149 Münster, Germany.
Chem Commun (Camb). 2019 Jan 24;55(9):1287-1290. doi: 10.1039/c8cc09204d.
Inteins change the structure and function of their host protein in a unique way and the Gp41-1 split intein is the fastest protein trans-splicing intein known to date. To design a photo-activatable variant, we have incorporated ortho-nitrobenzyl-tyrosine (ONBY) at the position of a structurally conserved phenylalanine in the Gp41-1-N fragment. Using irradiation at 365 nm, the splicing reaction was triggered with virtually unchanged rates. The partial cellular reduction of the nitro group in ONBY, previously observed during bacterial protein expression for several photo-caged amino acids, was overcome by periplasmatic expression and by using an E. coli K12(DE3) strain instead of BL21(DE3). Together, our findings provide new tools for the artificial photo-control of proteins.
内含子以独特的方式改变其宿主蛋白的结构和功能，而 Gp41-1 分裂内含子是迄今为止已知的最快的蛋白质转剪接内含子。为了设计光激活变体，我们在 Gp41-1-N 片段中结构保守的苯丙氨酸位置掺入了邻硝基苄基酪氨酸 (ONBY)。使用 365nm 的照射，拼接反应几乎以不变的速度触发。先前在几种光笼氨基酸的细菌蛋白表达过程中观察到 ONBY 的硝基部分的部分细胞还原，通过周质表达和使用大肠杆菌 K12(DE3)菌株而不是 BL21(DE3)克服了这一问题。总之，我们的发现为蛋白质的人工光控制提供了新的工具。
