Optimisation of engineered Escherichia coli biofilms for enzymatic biosynthesis of l-halotryptophans.
Optimisation of engineered Escherichia coli biofilms for enzymatic biosynthesis of l-halotryptophans.
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DOI：
10.1186/2191-0855-3-66
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发表时间：
2013-11-04
期刊：
影响因子：
3.7
通讯作者：
Overton TW
中科院分区：
文献类型：
--
作者：
Perni S;Hackett L;Goss RJ;Simmons MJ;Overton TW
由单一重组物种组成的工程生物膜作为新型生物催化剂在一系列应用中表现出显著的活性。在这项工作中，我们专注于利用大肠杆菌表达由质粒pSTB7编码的重组色氨酸合成酶，将5-卤吲哚转化为5-卤色氨酸，这是一种药物中间体。为了优化反应，我们比较了两种大肠杆菌K-12菌株（MC4100和MG1655）及其过量产生粘附素curli （PHL644和PHL628）的ompR234突变体。ompR234突变增加了MG1655和MC4100背景下的生物膜数量。在所有病例中，没有pSTB7质粒的细胞没有观察到5-卤代酚的转化。菌株PHL628 pSTB7和PHL644 pSTB7的工程生物膜比相应的浮游细胞产生更多的5-卤色氨酸。流式细胞术显示，绝大多数细胞在24小时的生物转化反应后仍然存活，无论是浮游形式还是生物膜形式，这表明细胞活力并不是生物膜反应性能提高的主要因素。对5-卤吲哚消耗、5-卤色氨酸合成和生物转化反应转化率的监测表明，菌株MG1655与MC4100之间、浮游细胞与生物膜细胞之间在色氨酸和吲哚代谢和转运方面存在内在差异。这项研究加强了在进行生物转化反应时彻底研究细菌生理学和做出明智的菌株选择的必要性。
Engineered biofilms comprising a single recombinant species have demonstrated remarkable activity as novel biocatalysts for a range of applications. In this work, we focused on the biotransformation of 5-haloindole into 5-halotryptophan, a pharmaceutical intermediate, using Escherichia coli expressing a recombinant tryptophan synthase enzyme encoded by plasmid pSTB7. To optimise the reaction we compared two E. coli K-12 strains (MC4100 and MG1655) and their ompR234 mutants, which overproduce the adhesin curli (PHL644 and PHL628). The ompR234 mutation increased the quantity of biofilm in both MG1655 and MC4100 backgrounds. In all cases, no conversion of 5-haloindoles was observed using cells without the pSTB7 plasmid. Engineered biofilms of strains PHL628 pSTB7 and PHL644 pSTB7 generated more 5-halotryptophan than their corresponding planktonic cells. Flow cytometry revealed that the vast majority of cells were alive after 24 hour biotransformation reactions, both in planktonic and biofilm forms, suggesting that cell viability was not a major factor in the greater performance of biofilm reactions. Monitoring 5-haloindole depletion, 5-halotryptophan synthesis and the percentage conversion of the biotransformation reaction suggested that there were inherent differences between strains MG1655 and MC4100, and between planktonic and biofilm cells, in terms of tryptophan and indole metabolism and transport. The study has reinforced the need to thoroughly investigate bacterial physiology and make informed strain selections when developing biotransformation reactions.
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影响因子：
11.4
作者：
Hu, Zhong-Ce;Zheng, Yu-Guo;Shen, Yin-Chu
通讯作者：
Shen, Yin-Chu
大肠杆菌K-12中生物膜开发的网络模型。
影响因子：
--
作者：
Shalá AA;Restrepo S;González Barrios AF
通讯作者：
González Barrios AF
影响因子：
11.4
作者：
Cortes-Lorenzo, C.;Rodriguez-Diaz, M.;Gonzalez-Lopez, J.
通讯作者：
Gonzalez-Lopez, J.
影响因子：
4.9
作者：
Goss, Rebecca J. M.;Newill, Philip L. A.
通讯作者：
Newill, Philip L. A.
影响因子：
3.2
作者：
Prigent-Combaret, C;Vidal, O;Lejeune, P
通讯作者：
Lejeune, P