The modification of heme special importer to improve the production of active hemoglobins in Escherichia coli.

IF 2 4区生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biotechnology Letters Pub Date : 2024-08-01 Epub Date: 2024-05-08 DOI:10.1007/s10529-024-03488-x

Zihan Zhang, Baodong Hu, Tao Zhang, Zhengshan Luo, Jingwen Zhou, Jianghua Li, Jian Chen, Guocheng Du, Xinrui Zhao

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Abstract

To enhance the import of heme for the production of active hemoproteins in Escherichia coli C41 (DE3) lacking the special heme import system, heme receptor ChuA from E. coli Nissle 1917 was modified through molecular docking and the other components (ChuTUV) for heme import was overexpressed, while heme import was tested through growth assay and heme sensor HS1 detection. A ChuA mutant G360K was selected, which could import 3.91 nM heme, compared with 2.92 nM of the wild-type ChuA. In addition, it presented that the expression of heme transporters ChuTUV was not necessary for heme import. Based on the modification of ChuA (G360K), the titer of human hemoglobin and the peroxidase activity of leghemoglobin reached 1.19 μg g^-1 DCW and 24.16 10³ U g^-1 DCW, compared with 1.09 μg g^-1 DCW and 21.56 10³ U g^-1 DCW of the wild-type ChuA, respectively. Heme import can be improved through the modification of heme receptor and the engineered strain with improved heme import has a potential to efficiently produce high-active hemoproteins.

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改造血红素特殊导入器，提高大肠杆菌活性血红蛋白的产量。

为了在缺乏特殊血红素输入系统的大肠杆菌C41（DE3）中增强血红素的输入以生产活性血蛋白，通过分子对接改造了大肠杆菌Nissle 1917的血红素受体ChuA，并过表达了血红素输入的其他元件（ChuTUV），同时通过生长试验和血红素传感器HS1检测血红素的输入。结果发现，ChuA 突变体 G360K 可导入 3.91 nM 的血红素，而野生型 ChuA 仅能导入 2.92 nM 的血红素。此外，研究还发现血红素转运体 ChuTUV 的表达并非血红素导入的必要条件。基于对 ChuA（G360K）的修饰，人血红蛋白的滴度和腿血红蛋白的过氧化物酶活性分别达到了 1.19 μg g-1 DCW 和 24.16 103 U g-1 DCW，而野生型 ChuA 的滴度和过氧化物酶活性分别为 1.09 μg g-1 DCW 和 21.56 103 U g-1 DCW。通过改造血红素受体可以改善血红素的导入，具有改善血红素导入能力的工程菌株有望高效生产高活性血蛋白。

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来源期刊

Biotechnology Letters 工程技术-生物工程与应用微生物

CiteScore

5.90

自引率

3.70%

发文量

108

审稿时长

1.2 months

期刊介绍： Biotechnology Letters is the world’s leading rapid-publication primary journal dedicated to biotechnology as a whole – that is to topics relating to actual or potential applications of biological reactions affected by microbial, plant or animal cells and biocatalysts derived from them. All relevant aspects of molecular biology, genetics and cell biochemistry, of process and reactor design, of pre- and post-treatment steps, and of manufacturing or service operations are therefore included. Contributions from industrial and academic laboratories are equally welcome. We also welcome contributions covering biotechnological aspects of regenerative medicine and biomaterials and also cancer biotechnology. Criteria for the acceptance of papers relate to our aim of publishing useful and informative results that will be of value to other workers in related fields. The emphasis is very much on novelty and immediacy in order to justify rapid publication of authors’ results. It should be noted, however, that we do not normally publish papers (but this is not absolute) that deal with unidentified consortia of microorganisms (e.g. as in activated sludge) as these results may not be easily reproducible in other laboratories. Papers describing the isolation and identification of microorganisms are not regarded as appropriate but such information can be appended as supporting information to a paper. Papers dealing with simple process development are usually considered to lack sufficient novelty or interest to warrant publication.