Team:ZJU-China/notebook.htm

This section is a summary of some outstanding ideas we developed during our brainstorming stage in the beginning two months after our team was founded.

List

Controlled chemistry plant

Synchronized bacterial clock

EPad

Methane alarm

Silkworm

Radio-communication

Plant doctor

Controlled chemistry plant

As we know, biofilms are always used for chemical transformations in biorefineries. To utilize biofilms more efficiently, we need to control them and make them replaced. Here we expect to use dispersal proteins along with population-driven quorum-sensing switches to accomplish this goal.

We found some paper on related studies. In one of them, the researchers form an initial colonizer biofilm in a microfluidic device, introduce a second cell type called disperser into this existing biofilm, form arobust dual-specied biofilm and displace the initial colonizer biofilm with an extraceller signal from the disperser cells. They also remove the disperser biofilm with a chemically induced switch, and the consortial population could tune. The sketch followed show the mechanism of it.

The two E.coli cell types communicate by using the LasI/LasR QS module of P.aeruginosa. In the disperser cell, the LasI protein(autoinducer synthase) is constitutively produces and synthesizes the QS signal 3oC12HSL. 3oC12HSL freely diffuses into the initial colonizer cell and makes a complex with LasR(LuxR family transcriptional regulator), and the 3oC12HSL+LasR complex induces biofilm dispersal protein BdcAE50Q by activating the lasI promoter. BdcAE50Q disperses biofilms by binding cyclic diguanylate. The biofilm dispersal protein Hha13D6 in the disperser cell is induced upon adding IPTG. Hha13D6 disperses biofilms by activating proteases.

(From "Synthetic quorum-sensing circuit to control consortial biofilm formation and dispersal in a microfluidic device")

We want to develop the system and found a system contains three E.coli cell types, in which the first one can be replaced by the second one, the second one can be replaced by the third one and the third one can be replaced when added some chemical signals. Thus, this kind of controlled chemistry plant can be applied to a three-step chemical reaction and each kind of cells will finish one of them in sequence, just as follows.

Reference:

1. "Synthetic quorum-sensing circuit to control consortial biofilm formation and dispersal in a microfluidic device"

(http://www.nature.com/ncomms/journal/v3/n1/full/ncomms1616.html)

2. "Engineering microbial consortia: a new frontier in synthetic biology"

(http://www.ncbi.nlm.nih.gov/pubmed/18675483)

3. "Applications of quorum sensing in biotechnology"

(http://www.springerlink.com/content/j16470w7g3838851/)

4. "A synthetic Escherichia coli predator-prey ecosystem"

(http://www.nature.com/msb/journal/v4/n1/full/msb200824.html)

Synchronized bacterial clock

Human orchestrate their activities in different places because of their synchronizing clocks. Isn't it cool if bacteria can coordinate their molecular timepieces in the same way as we human do?

We found that Hasty and colleagues have constructed a network of genes and proteins in E.coli that acts as a molecular clock and can be synchronized across a population of the bacteria. The mechanism can be described briefly as follows.

A promoter(PluxI)drives the production of LuxI, an enzyme that synthesizes the quorumsensing signal acyl-homoserine lactone (AHL). Another PluxI control ths the production of AiiA, a protein that catalyses the degradation of AHL, and a third PluxI triggers the synthesis of a variant of green fluorescent protein called yemGFP. An AHL receptor, LuxR, is constitutively expressed. The authors combined these components to form an autoinducng circuit (AHL activates LuxR, and the AHL-LuxR complex induces PluxI-driven luxI transcription and yemGFP production) with a time-delayed negative feedback loop (the AHL-LuxR complex induces PluxI-driven production of AiiA, which degrades AHL). The result was a population of bacteria that produce synchronized pulses of fluorescence, coordinated by quorum sensing.

(From "Synthetic biology: synchronized bacterial clock")

We expect to apply the network to medical field. Since the network can make an oscillation, it can be used to release medicine periodically. What is more, we wish to add another sensor in the network. The sensor can sense a signal which is released because of a certain disease, and thus the cells can sense the signal and start the oscillation which releases the medicine periodically. For instance, when a man hasdiabetes, the high density blood sugar will start an insulin-releasing oscillation.

Reference:

1. "Synthetic biology: synchronized bacterial clock"

(http://www.nature.com/nature/journal/v463/n7279/full/463301a.html)

2. "A synchronized quorum of genetic clock"

(http://www.nature.com/nature/journal/v463/n7279/abs/nature08753.html)

3. "bacteria collaborate to sense arsenic"

(http://www.nature.com/nature/journal/v481/n7379/full/481033a.html)

EPad

Ipad is popular all over the world, but how about an Epad? Have you thought that there is a tablet whose screen is made up of E.coli and it can sense your touch to display different colors?

MscL is a kind of membrane protein of E.coli. They control the transportation of matter, but they are usually closed. When the balance between intracellular and extracellular is broken, or the cytomembrane senses some unusual mechanical force, they will open and let the pressure disappear. We hope to utilize the protein and found a network which can display fluorescence of different color according to the state of MscL. In this way, a pressure-sensing and -controlling screen made of E.coli can be built.

Reference:

1. "Release of content through mechano-sensitive gates in pressurized liposomes"

(http://www.pnas.org/content/107/46/19856.short)

2. "Mechanical force and cytoplasmic Ca2+ activate yeast TRPY1 in parallel"

(http://www.springerlink.com/content/xg2q0601j73q1r31/)

Methane alarm

The concentration of methane in the coal mine is an important indicator which affects the life safety of miners. There should be alarm when the concentration of methane is between 1.0-1.5percent in the coal mine under the Chinese law. We aim to design a methane alarm for coal mine by methanotrophs, which consists of four parts: decomposition, sensor, signal tuner and output. Decomposition part takes in methane and conducts catalytic decomposition of methane to CO2 or assimilates methane as its own organic compounds. This function is the naturally exist in methanotrophs. Sensor senses the concentration of methane and produces activator A. As for signal tuner part, we plan to find a substance B which is sensitive to the concentration of activator A. Activator A and substance B combines to control the activity of substance C. Substance C acts as the switch of promoter--when the concentration of methane has been exceeded the alarm threshold, promoter will turn on. Output part will give the alarm by producing pigment of red lycopene when promoter is open. We plan to put methanotrophs on the miner lamp. When the bacteria produce pigment, the light through bacteria will turn out to be red.

Reference:

1. "Biological Methane Oxidation: Regulation, Biochemistry, and Active Site Structure of Particulate Methane Monooxygenase"

(http://informahealthcare.com/doi/full/10.1080/10409230490475507)

2. "Importance of methane-oxidizing bacteria in the methane budget as revealed by the use of a specific inhibitor"

(http://www.nature.com/nature/journal/v356/n6368/abs/356421a0.html)

Silkworm

Hangzhou is famous for silk. We wish to let engineered bacterial produce silk proteins, just like lovely silkworm. A machine which can make threadlike things with the proteins will be used to make silk.

Reference:

1. "Morphology and primary crystal structure of a silk-like protein polymer synthesized by genetically engineered Escherichia coli bacteria"

(http://onlinelibrary.wiley.com/doi/10.1002/bip.360340808/abstract)

2. "Biotechnological Production of Spider-Silk Proteins Enables New Applications"

(http://onlinelibrary.wiley.com/doi/10.1002/mabi.200600255/full)

Radio-communication

Synthetic biology always needs bacteria to sense an input signal and output another. It occurs to us that we can use two kinds of bacteria to accomplish the task. The first one kind of bacteria senses the input signal and then fluoresces, while the second one senses the fluorescence from a distance far away from the first one and then output the final signal. It do not need a straight contact between the two kinds of bacteria, seeming like a radio-communication.

Plant doctor

When the leaves of some kinds of plants are bitted by insects, the substance A in the juice of plants will react with the one B in the spit of the insects. As a consequence, a volatile matter is produced, which attract the natural enemy of the insect. We plan to engineer the bacteria, having a symbiosis with plants, with genes of the enzymes which catalyze the production of the substance A in the juice. The bacteria will make plant doctors.