Abstract：
Transcriptional regulation is of great significance for cells to maintain homeostasis and, meanwhile, represents an innovative but less explored means to control biological processes in synthetic biology and bioengineering. Herein we devised a T7 RNA polymerase (T7RNAP) variant through replacing an essential lysine located in the catalytic core (K631) with Nε-acetyl-l-lysine (AcK) via genetic code expansion. This T7RNAP variant requires the deacetylase activity of NAD-dependent sirtuins to recover its enzymatic activities and thereby sustains sirtuin-dependent transcription of the gene of interest in live cells including bacteria and mammalian cells as well as in in vitro systems. This T7RNAP variant could link gene transcription to sirtuin expression and NAD availability, thus holding promise to support some relevant research.
摘要：
转录调控对细胞维持稳态具有重要意义,同时,代表了一种创新但较少探索的方法来控制合成生物学和生物工程中的生物过程。在此，我们设计了一种T7RNA聚合酶（T7RNAP）变体，通过遗传密码扩展，将位于催化核心（K631）中的必需赖氨酸替换为Nε-乙酰基-1-赖氨酸（AcK）。该T7RNAP变体需要NAD依赖性沉默调节蛋白的脱乙酰酶活性以恢复其酶活性,从而在活细胞(包括细菌和哺乳动物细胞)以及体外系统中维持目的基因的沉默调节蛋白依赖性转录。这种T7RNAP变体可以将基因转录与沉默酶表达和NAD可用性联系起来，因此，有希望支持一些相关研究。
