IGEM:Stanford/2009/Groups/Team Isis

Brita: New and Improved

• Brief recap/outline: We've divided up the idea as follows:
  • 1. Indication and Quorum sensing (Leon)
    • Because we don't like dirty catheters
    • A good old estrogen receptor we could use
    • I'm just going to post this here so i don't forget about it later
      • I'm thinking we could put GFP downstream of the code for this receptor so that the GFP is turned off when estrogen is present]
  • 2. Metabolism and Degradation (Isis)
    • Biodegradation of bisphenol A by cells and cell lysate from Sphingomonas sp. strain AO1.
    • Purification of Cytochrome P450 and Ferredoxin, Involved in Bisphenol A Degradation, from Sphingomonas sp. Strain AO1
    • Enzymatic degradation of alkylphenols, bisphenol A, synthetic estrogen and phthalic ester
  • 3. Apotosis (Ariana)

Ideas

• using luciferase to produce light for photosynthesis
• inject chloroplasts into fungus?
• controlling quorum sensing with riboswitch
• interspecies quorum sensing
• sensing/removing environmental estrogens - bioremediation

Questions

Quorum Sensing

• Bacteria responding to different concentrations on inducers

Biofuels

• Can a chemostat be used to culture biofuel producing organisms?
• How are biofuels transported out the cells and how can this be improved?
• What genes can be pared away/controlled so the cell uses the maximizes energy usage for fuel output?
• Can light adsorption range be improved by expressing other photosensitive pigments?

Photosynthesis

• Questions to think about:
  • What organism do we want to use? What pigments does it express? What pigments would expand its photosynthetic range?
  • What are the natural biosynthesis pathways of these pigments? Are there overlaps between the endogenous pigment pathway and the pigment pathway we want to express?

--Isis

• Interesting general information sites:
  • Review of Photosynthesis -Isis
  • Overview of Photosynthetic Pigments -Isis
  • Photosynthetic Pigments: Structure and Spectroscopy - Isis

• Interesting photosynthetic pigment biosynthesis papers:
  • Chlorophylls
  • Carotenoids
    • Molecular genetics of the carotenoid biosynthesis pathway in plants and algae
    • Eubacteria show their true colors: genetics of carotenoid pigment biosynthesis from microbes to plants
  • Phycobilins
    • Biosynthesis of phycobilins
    • Photosynthetic Pigments: Perplexing Persistent Prevalence of ‘Superfluous’ Pigment Production
  • Multiple pigments
    • Genes Involved in the Biosynthesis of Photosynthetic Pigments in the Purple Sulfur Photosynthetic Bacterium Thiocapsa roseopersicina
    • Efficient Phage-Mediated Pigment Biosynthesis in Oceanic Cyanobacteria

• Interesting metabolic engineering of pigment pathways papers:
  • Chlorophyll b Expressed in Cyanobacteria Functions as a Light-harvesting Antenna in Photosystem I through Flexibility of the Proteins
  • Metabolic engineering of carotenoid biosynthesis in plants
  • Molecular breeding of carotenoid biosynthetic pathways
  • Metabolic Engineering of Plant Secondary Metabolism
  • Sheer Madness-Photosynthetic E coli

• Other iGEM projects:
  • Towards a bacterial assembly line

• Reprint of stuff I said on the Ideas page:

1. First, to make chlorophyll absorb different wavelengths, we would have to modify the chemical structure which conveniently has already been done by nature. There are a bunch of photosynthetic pigments that cover most of the spectrum.
2. I think what would be interesting would be to try to express these pigments in not-native hosts. Like have a green algae express a red algae pigment thereby expanding its photosynthetic range (?) (note: expression of the pigment doesn't really mean it will increase photosynthesis because it would have to link in with the endogenous photosynthetic machinery... its hard to tell if that would happen on its own or not.)
3. Major problem: the pathways to synthesize these molecules are complicated and require multiple enzymatic steps. Take chlorophyll biosynthesis for example. This makes it really hard to just transfer the pathway into a new host
4. (presentation idea?!) It might be interesting to see if there is much overlap between the biosynthetic pathways of the different pigments... that way we may only need to introduce a few enzymes to get pigment expression. It is also important to check if the enzymes are even clonable

--Isis

Thanks for posting all this stuff on the wiki, Isis! The chapter that you posted about chlorophyll structure and spectroscopy makes it seem like it would be really difficult to express a particular chlorophyll in a different organism; it talked about 17 different enzymatic steps for the production of chlorophyll a. Or is there a lot of crossover/common enzymes between pathways for different chlorophylls? I couldn't tell from what I've read so far, but maybe I just missed it. Also, how would we go about making it compatible with the host organism? Would we introduce it as a vector or try to slide it into the nuclear or chloroplast chromosome? I guess if we did either of the first two, we'd have to find a way to make sure that it gets targeted to the correct plasma membrane of the chloroplast. I saw some articles about scientists finding patterns in the sequences of proteins inside the nuclear chromosomes that do get targeted for relocation to the chlorophyll, so maybe we could use that.

On another note, would it be possible to encode luciferase into photosynthetic bacteria or plant cells and then have it relocate to the photosynthetic machinery and be a source of light for photosynthesis during night? Luciferase still glows at night, right? And then perhaps we could add a riboswitch so that we could downregulate it during the day. Probably not at all feasible, but just a thought.... -Ariana

Plastic Degredation

• How do fungi degrade stuff?
  • Fungi Online - all you ever wanted to know about fungi -Isis
  • Interesting fungus degrading-stuff papers:
    • Fungal Communities Associated with Degradation of Polyester Polyurethane in Soil - Isis
    • Purification and characterization of a biodegradable plastic-degrading enzyme from Aspergillus oryzae -Isis
    • Purification and Properties of a Polyester Polyurethane-Degrading Enzyme from Comamonas acidovorans TB-35 -Isis
• How are fungi cultured/are they genetically tractable?
  • Transforming Fungus
• What are the degradation products of plastics?
• How can degradation be controlled? Riboswitches? A kill switch?
• Other iGEM projects:
  • Duke 2008 - The Duke team degraded polyethylene by expressing an alkane degradation pathway in E.coli - CVL

Other Stuff

• What is the mechanism of quorum sensing?
  • From wikipedia: "Bacteria that use quorum sensing constantly produce and secrete certain signaling molecules (called autoinducers or pheromones). These bacteria also have a receptor that can specifically detect the signaling molecule (inducer). When the inducer binds the receptor, it activates transcription of certain genes, including those for inducer synthesis. There is a low likelihood of a bacterium detecting its own secreted inducer. Thus, in order for gene transcription to be activated, the cell must encounter signaling molecules secreted by other cells in its environment. When only a few other bacteria of the same kind are in the vicinity, diffusion reduces the concentration of the inducer in the surrounding medium to almost zero, so the bacteria produce little inducer. However, as the population grows the concentration of the inducer passes a threshold, causing more inducer to be synthesized. This forms a positive feedback loop, and the receptor becomes fully activated. Activation of the receptor induces the up regulation of other specific genes, causing all of the cells to begin transcription at approximately the same time. This coordinated behavior of bacterial cells can be useful in a variety of situations. For instance, the bioluminescent luciferase produced by V. fischeri would not be visible if it were produced by a single cell. By using quorum sensing to limit the production of luciferase to situations when cell populations are large, V. fischeri cells are able to avoid wasting energy on the production of useless products."

Past Meetings

• What is the natural function of hammerhead ribozymes?
  • They are found in viroids, which are plant viruses that have a naked RNA genome (no protein coat). To protect the genome from degradation by ribonucleases it is packed very tightly until it enters the host. Once it gets in, the hammerhead ribozyme part is activated to cleave the genome open so it an be translated.
• Next meeting: Thursday at 5pm. Location TBD.
  • Agenda: Discuss RNA paper.

• Things to think about/research:
  • How to read/write DNA
  • Using cells as sensors? How to Input/Output information? How to control behavior?

Storing info in DNA

• Here's an article about storing "It's a small world" in DNA which Ariana mentioned at our meeting.
  • Deinococcus radiodurans

Questions from outsiders

• Robert 21:45, 7 March 2009 (EST):I am really interested in the engineered photosynthetic magical machines. If we do introduce a new pathway, will this theoretically increase efficiency of energy-gathering from the cell? How would we measure output of energy or gather this energy? I don't know too much about P.S. - sorry for the elementary questions!