A Non-natural Protein Rescues Cells Deleted for a Key Enzyme in Central Metabolism
A Non-natural Protein Rescues Cells Deleted for a Key Enzyme in Central Metabolism
复制标题
DOI：
10.1021/acssynbio.6b00336
复制
发表时间：
2017-04-01
影响因子：
4.7
通讯作者：
Hecht, Michael H.
中科院分区：
文献类型：
--
作者：
Digianantonio, Katherine M.;Korolev, Maria;Hecht, Michael H.
合成生物学的一个重要目标是创造新的蛋白质，在生物体中提供维持生命的功能。最近的尝试，以产生新的蛋白质主要集中在合理的设计，涉及大量的计算工作。相比之下，大自然并不是先验地设计序列。相反，大自然依赖达尔文进化论从非设计的序列空间中选择具有生物功能的序列。为了在实验室中模拟自然选择，我们梳理了新序列的文库，并选择了拯救E。大肠杆菌细胞中条件必需基因的缺失。一个这样的基因，gltA，编码柠檬酸合酶，负责代谢进入柠檬酸循环的酶。新生蛋白SynGItA作为Ag/tA的拯救者被分离。然而，SynGRA不是一种酶。相反，SynGItA允许细胞通过改变替代代谢途径的调节来从中心碳和能量代谢的缺陷中恢复。具体而言，SynG 1 tA显著增强了丙酸降解途径中编码甲基柠檬酸合酶的基因prpC的表达。这种内源性蛋白质具有混杂的催化活性，当过表达时，其补偿柠檬酸合酶的缺失。虽然负责这种过度表达的分子细节尚未阐明，但结果清楚地表明，与自然界中的序列无关的非天然蛋白质可以通过改变天然生物体中的基因调控来提供维持生命的功能。
An important goal of synthetic biology is to create novel proteins that provide life-sustaining functions in living organisms. Recent attempts to produce novel proteins have focused largely on rational design involving significant computational efforts. In contrast, nature does not design sequences a priori. Instead, nature relies on Darwinian evolution to select biologically functional sequences from nondesigned sequence space. To mimic natural selection in the laboratory, we combed through libraries of novel sequences and selected proteins that rescue E. coli cells deleted for conditionally essential genes. One such gene, gltA, encodes citrate synthase, the enzyme responsible for metabolic entry into the citric acid cycle. The de novo protein SynGItA was isolated as a rescuer of Ag/tA. However, SynGRA is not an enzyme. Instead, SynGItA allows cells to recover from a defect in central carbon and energy metabolism by altering the regulation of an alternative metabolic pathway. Specifically, SynG1tA dramatically enhances the expression of prpC, a gene encoding methylcitrate synthase in the propionate degradation pathway. This endogenous protein has promiscuous catalytic activity, which when overexpressed, compensates for the deletion of citrate synthase. While the molecular details responsible for this overexpression have not been elucidated, the results clearly demonstrate that non-natural proteins unrelated to sequences in nature can provide life-sustaining functions by altering gene regulation in natural organisms.