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Engineered single- and multi-cell chemotaxis pathways in E. coli.

ABSTRACT: We have engineered the chemotaxis system of Escherichia coli to respond to molecules that are not attractants for wild-type cells. The system depends on an artificially introduced enzymatic activity that converts the target molecule into a ligand for an E. coli chemoreceptor, thereby enabling the cells to respond to the new attractant. Two systems were designed, and both showed robust chemotactic responses in semisolid and liquid media. The first incorporates an asparaginase enzyme and the native E. coli aspartate receptor to produce a response to asparagine; the second uses penicillin acylase and an engineered chemoreceptor for phenylacetic acid to produce a response to phenylacetyl glycine. In addition, by taking advantage of a 'hitchhiker' effect in which cells producing the ligand can induce chemotaxis of neighboring cells lacking enzymatic activity, we were able to design a more complex system that functions as a simple microbial consortium. The result effectively introduces a logical 'AND' into the system so that the population only swims towards the combined gradients of two attractants.

SUBMITTER: Goldberg SD

PROVIDER: S-EPMC2710872 | biostudies-other | 2009

REPOSITORIES: biostudies-other

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Engineered single- and multi-cell chemotaxis pathways in E. coli.

Goldberg Shalom D SD Derr Paige P DeGrado William F WF Goulian Mark M

Molecular systems biology 20090616

We have engineered the chemotaxis system of Escherichia coli to respond to molecules that are not attractants for wild-type cells. The system depends on an artificially introduced enzymatic activity that converts the target molecule into a ligand for an E. coli chemoreceptor, thereby enabling the cells to respond to the new attractant. Two systems were designed, and both showed robust chemotactic responses in semisolid and liquid media. The first incorporates an asparaginase enzyme and the nativ ...[more]

PMID: 19536206

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Project description:BACKGROUND: ?-carotene is a carotenoid compound that has been widely used not only in the industrial production of pharmaceuticals but also as nutraceuticals, animal feed additives, functional cosmetics, and food colorants. Currently, more than 90% of commercial ?-carotene is produced by chemical synthesis. Due to the growing public concern over food safety, the use of chemically synthesized ?-carotene as food additives or functional cosmetic agents has been severely controlled in recent years. This has reignited the enthusiasm for seeking natural ?-carotene in large-scale fermentative production by microorganisms. RESULTS: To increase ?-carotene production by improving the isopentenyl pyrophosphate (IPP) and geranyl diphospate (GPP) concentration in the cell, the optimized MEP (methylerythritol 4-phosphate) pathway containing 1-deoxy-D-xylulose-5-phosphate synthase (DXS) and isopentenyl pyrophosphate isomerase (FNI) from Bacillus subtilis, geranyl diphosphate synthase (GPPS2) from Abies grandis have been co-expressed in an engineered E. coli strain. To further enhance the production of ?-carotene, the hybrid MVA (mevalonate) pathway has been introduced into an engineered E. coli strain, co-expressed with the optimized MEP pathway and GPPS2. The final genetically modified strain, YJM49, can accumulate 122.4±6.2 mg/L ?-carotene in flask culture, approximately 113-fold and 1.7 times greater than strain YJM39, which carries the native MEP pathway, and YJM45, which harbors the MVA pathway and the native MEP pathway, respectively. Subsequently, the fermentation process was optimized to enhance ?-carotene production with a maximum titer of 256.8±10.4 mg/L. Finally, the fed-batch fermentation of ?-carotene was evaluated using the optimized culture conditions. After induction for 56 h, the final engineered strain YJM49 accumulated 3.2 g/L ?-carotene with a volumetric productivity of 0.37 mg/(L?·?h?·?OD600) in aerobic fed-batch fermentation, and the conversion efficiency of glycerol to ?-carotene (gram to gram) reached 2.76%. CONCLUSIONS: In this paper, by using metabolic engineering techniques, the more efficient biosynthetic pathway of ?-carotene was successfully assembled in E. coli BL21(DE3) with the optimized MEP (methylerythritol 4-phosphate) pathway, the gene for GPPS2 from Abies grandis, the hybrid MVA (mevalonate) pathway and ?-carotene synthesis genes from Erwinia herbicola.

Dataset Information

Engineered single- and multi-cell chemotaxis pathways in E. coli.

Publications

Engineered single- and multi-cell chemotaxis pathways in E. coli.

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