Overview

BUCT's project is designed to help people improve their scalp microecology. We know the negative effects of non-esterified fatty acids produced by the breakdown of sebum by microorganisms such as Malassezia on the scalp. Therefore, after considering many comprehensive factors, we believe that E. coli Nissle 1917 is a safe, easy to operate and easy to absorb non-esterified fatty acids chassis.

In order to make the design conform to the prototype of scalp care products, safety, efficacy and acceptability were fully considered when selecting the micromolecule used for synthesis. Caffeine, a natural compound found in plants, has been shown to benefit hair follicles and promote healthy hair. 2-phenylethanol (2-PE) is an important perfume compound with rose fragrance, which is widely used in cosmetics, perfume, food and other industries.

Part-1 Enhanced consumption of fatty acids

The fadR of E. coli has been shown to play a dual role in transcription of the genes of bacterial fatty acid metabolism. The FadR protein acts as a repressor of β-oxidation. FadR also acts as a sensor of fatty acid availability in the environment. FadR DNA binding is antagonized by long chain acyl-CoAs. Therefore, knockdown of fadR can enhance the oxidative decomposition of fatty acids.

BUCT knocked out fadR once in 2021, but we had to knock out fadR again in E. coli Nissle 1917 because the original strain was discarded during laboratory cleaning. But we saved the information that we used to knock out fadR, and we knocked it out of the genome again.

Part-2 Synthesis of Caffeine

The main biosynthetic pathway of caffeine is a sequence consisting of xanthine → 3-methylxanthine → theobromine → caffeine. In three of the four reactions, the gene encoding N-methyltransferase was involved.

The gene of TcAncCS1 is derived from Cocobroma. Xanthine can be converted to 3-methylxanthine (3-mX) by TcAncCS1 from Theobroma. TcCS2 from C. sinensis has a higher propensity to recognize 3-mX and can convert 3-mX to theobromine.

Part-3 Synthesis of 2-Phenylethyl Alcohol

In nature, 2-PE is found in a variety of plants, including apples, almonds, bananas, roses, hyacinths, jasmine, and lilies. It can be used as edible spices, daily chemical products fragrance, can also modulate rose fragrance, jasmine fragrance, clove fragrance and other flower fragrance, used in the production of soap, shower gel, cosmetics and other products.

BUCT hopes to use 2-PE as an example to give E. coli Nissle 1917 the ability to sweetener hair in addition to improving scalp microecology. We use the natural phenylalanine synthesis pathway of E. coli to synthesize 2-PE. aroG was used to enhance the accumulation of a key intermediate, DAHP. To allow the production to flow towards 2-PE, we also used aro10 (encoding 2-keto acid decarboxylase) and adh6 (encoding phosphate reductase) from Saccharomyces cerevisiae.

After the first 2-PE synthesis, we compared the results with JLU-China and improved the design.

tyrB enables our probiotics to utilize phenylalanine, so we will add phenylalanine to the substrate to assist 2-PE synthesis. adh1 and adh6 are isoenzymes, and aro10 and kdcA are isoenzymes. We'll use different enzymes from both teams to compare conditions and choose the best combination.

Part-4 Suicide system

To reduce strain instability under natural conditions, we hoped to control the competitive advantage of E. coli Nissle 1917 by quorum sensing. Through modeling and calculation, Nissle can control the concentration as much as possible within the range specified by us on the premise of satisfying the function. (see: )