This project includes two parts: a) to form and to destroy the anaerobic environment b) power-generating by anaerobic fermentation

a.The directed motion of E.coli

In normal condition, the motion direction of flagella of E.coli is clockwise which results in the disorderly movement. In our circuit (see Fig.1), promoter PuerI converts the chemical signal (the concentration of H+) into biochemical signal (AHL), changing the motion direction of flagella to be anticlockwise and thus the motion of E.coli becomes orderly towards one direction.

Fig.1 After sensing the H+ in the solution, PureI expresses protein AHL after several steps which can combine with the membrane receptor expressed downstream. Then the motion direction of flagella can be changed.

b.To form the anaerobic environment

The output of electricity is achieved by catalyzing the substrate in anaerobic environment which is formed by biofilm produced by the constellation of E.coli. Thus, the formation of anaerobic environment is actually the formation of biofilm. The expression of gene CsgD can control the production of biofilm positively. Conclusively, we can produce biofilm by utilizing the expression of CsgD.
As the biofilm is formed, the contact between bacterium becomes stronger, which leads to the decrease of oxygen concentration. Now, the promoter Pvgd which senses the concentration of oxygen is activated and also activates the expression of CsgD.(see Fig.2) Gradually, the anaerobic environment is formed.

Fig.2 The bacteria produce CsgD to form biofilm after sensing H+, which promotes the form of anaerobic environment. Then the promoter sensing the concentration of oxygen is activated to strengthen the same effect of forming biofilm.

c.power-generating by anaerobic fermentation

The final goal of our project is to generate electricity. Therefore, we design a circuit (see Fig.3) with 3 enzymes, including pflB, fdhF, and NOX, to complete the circuit of electricity generation. ( see Fig.4)

Fig.3 The protein expressed by PfdhF inhibits downstream protein which will inhibit target proteins. That means, in this circuit, the protein we need, pflB, fdhF, and NOX, will be expressed and lead to electricity generation.

Fig.4 Pyruvate is the products of the reactions of glycolysis and also the material of generating electricity. After the catalysis of the 3 kind of enzymes we used, pflB, fdhF, and NOX, pyruvate can produce electron which react in the electrode reaction.

d.control of the density of bacterium

We use CcdB which is toxic to cells to control the density of E.coli. When the cell population grows, the concentration of AHL will grow to a specific scale and activate the expression of CcdB to kill some of the E.coli. (see Fig.5)
Fig.5
We also a desigh a safety device with which we can make cells commit suiside when necessary. (see Fig.6)
Fig.6