Functional replacement of histidine in proteins to generate noncanonical amino acid dependent organisms,Journal of the American Chemical Society

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Functional replacement of histidine in proteins to generate noncanonical amino acid dependent organisms
Journal of the American Chemical Society ( IF 14.4 ) Pub Date : 2018-03-06 , DOI: 10.1021/jacs.7b13452
Fei Gan ₁ , Renhe Liu ₁ , Feng Wang ₁ , Peter G. Schultz _{1,

2}

Affiliation

Simple strategies to produce organisms whose growth is strictly dependent on the presence of a noncanonical amino acid are useful for the generation of live vaccines and the biological containment of recombinant organisms. To this end, we report an approach based on genetically replacing key histidine (His) residues in essential proteins with functional His analogs. We demonstrate that 3-methyl-l-histidine (MeH) functionally substitutes for a key metal binding ligand, H264, in the zinc-containing metalloenzyme mannose-6-phosphate isomerase (ManA). An evolved variant, Opt5, harboring both N262S and H264MeH substitutions exhibited comparable activities to wild type ManA. An engineered Escherichia coli strain containing the ManA variant Opt5 was strictly dependent on MeH for growth with an extremely low reversion rate. This straightforward strategy should be applicable to other metallo- or nonmetalloproteins that contain essential His residues.

中文翻译：

蛋白质中组氨酸的功能性置换以产生非规范氨基酸依赖性生物

生产其生长严格依赖于非经典氨基酸存在的生物体的简单策略可用于生产活疫苗和重组生物体的生物控制。为此，我们报告了一种基于用功能性 His 类似物基因替换必需蛋白质中的关键组氨酸 (His) 残基的方法。我们证明 3-甲基-l-组氨酸 (MeH) 在功能上替代了含锌金属酶甘露糖 6-磷酸异构酶 (ManA) 中的关键金属结合配体 H264。进化变体 Opt5 具有 N262S 和 H264MeH 替代，表现出与野生型 ManA 相当的活性。含有 ManA 变体 Opt5 的工程大肠杆菌菌株严格依赖于 MeH 以极低的回复率生长。

更新日期：2018-03-06

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