User:Kiran S. Yemul

Environmental Group: Treehuggers Inc.



 * Kiran S. Yemul
 * Course 20, MIT
 * Email me through OpenWetWare

Team Members: Benji Moncivaiz, Paul Youchak, Kiran Yemul

Team Leaders: Roshini Zacharia, Neelima Yeddanapudi

Challenge: To find ways to lessen the impact of human byproducts or technology on the environments.

Ideas:

Organisms that Metabolize Ionizing Radiation from Nuclear wastes. In an effort to create safer and more developed ways to dispose of Nuclear waste, organisms have been found which can use the radiation emitted and naturally live off of it. In high enough numbers these organism might be able to lower the effect of wastes on the environment making it easier to dispose of wastes.

A membrane which can filter larger materials making cleaner water for third world countries. Biological organisms to make membrane, or actually use membranes very similar to human, one which is permeable to water but not other contaminants. Propose a set of materials which can degrade and breakdown organic material wastes in third world countries so as to prevent harmful means of discard by the local populace.

Organic Solar Panel creation. A solar panel made of living organism can have great effects on removing Carbon Dioxide from the atmosphere. Placing these organisms in a location where there is great quantities of sunlight would be great, however keeping such organisms alive in this harsh environment would be expensive and problematic. If we can store some of the energy they create from the sun then we could make this proposal economically viable to technological photovoltaics but at the same time removing Carbon from atmosphere.

Chemical Water Pollution. Find way to locate and remove chemicals from the water. Organism which natural use chemicals can be employed, either through actual presence or by DNA manipulation.

Project Development Notebook
This is the format by which I've kept my project development notebook over the past two months. The Project Log section contains my summary of work done on class days, however, a great deal of at-home background research lead to the questions and conclusions presented in each day's log.

Three Ideas Presentation

 * 1) Photovoltaics - Organic Solar Panels: http://www.solarserver.de/solarmagazin/solar-report_0807_e.html
 * 2) Nuclear Waste Eaters: Bacteria that thrive on nuclear waste could be used for better disposal
 * 3) Aquaporin Filtering Project: A membrane that could be produce cheap for water filtration
 * 4) Better Garbage Disposal brainstorming - *As of our meeting on Sunday, 3/9*, we have decided to scrap this idea for our Three Ideas Presentation.  Our group consensus is that the other ideas present more interesting engineering challenges to us.

Organic Photovoltaics
Berkeley: http://www.lbl.gov/tech-transfer/techs/lbnl2338.html
 * 1) 1 One issue with photovoltaic cells is absorption/emission on the edges of the spectrum over which energy can be harvested by these organic materials. More study needs to be done to see how this is dealt with in the bacterial cell.  In the meantime, for the purposes of this project, this presentation has graphs comparing the compounds used with optimal wavelengths.  Interesting project here. http://www.princeton.edu/~pccm/outreach/REU2005/REU2005Presentations/gerardi.pdf
 * 2) 2 How Photosynthetic Bacteria Harvest Solar Energy: http://jb.asm.org/cgi/content/full/181/13/3869
 * 3) 3 Good diagram of an organic photovoltaic cell structure: http://books.google.com/books?id=5PMa13y44FYC&pg=PA405&lpg=PA405&dq=organic+photovoltaics+wavelength&source=web&ots=u844zvbJCo&sig=Qcdr0qGdw1WJ25ZcxtKwkmLaAhI&hl=en#PPA406,M1
 * 4) 4 Wow, Sigma-Aldrich has a list of materials in it's catalog that could be used in photovoltaic cells. Catagories include: dyes, conducting materials, and nanoparticles.  Possible project goal:  What is the best combination of dyes and conducting materials to get maximum energy from our cell? http://www.sigmaaldrich.com/catalog/search/TablePage/19352445
 * 5) 5 What has been found so far in terms of comparing efficiency of different dyes? Different conducting materials? Efficient Organic Dyes containing Benzofuran: http://pubs.acs.org/cgi-bin/abstract.cgi/joceah/2007/72/i10/abs/jo0625150.html

Project Log
Filtering Membranes: Cleaning organisms for current water filters – cheap solutions to h2o contamination in 3rd world. What gets stuck in the filters? Organic particles, bacteria/microorganisms, Biological filter system possibilities: diffusion, channel proteins - aquaporins, Production How to control growth of membrane? How will it be safe for humans to use?
 * March 4, 2008:

Organic Solar Panels: How will energy be harvested? In what form?

Things to be researched: Radiation eaters What levels of radiation are we dealing with? Spectrum, intensity -Capabilities of fungi, a realistic amount of fungi -	nutrients for fungi -	what do they absorb, what’s emitted?

Filtering project: Aquaporins and their capabilities How to create actual physical membrane structure – size and scale. Other projects that have dealt with aquaporins  Organic Solar Panel How to harvest – mechanically, or chemical – glucose, electrical What other projects have been done? How to improve current photovoltaic cell projects using biological methods  Garbage Disposal brainstorming Sewage inappropriately disposed of Containers – speed up the natural biodegradation

Tried to gain better of standing of what/how photovoltaics work, how is the energy harvested, and how do organics differ from inorganics. The main issue with our initial idea is that we wanted to use whole, live bacteria to generate the electron transfer. This is going to require an input of nutrients, medium, etc. to keep the solar cell running. Today's organic cells simply use the donor/acceptor molecules from the bacteria but do not require a whole, functional cell. Which is why they are so much better overall than current inorganic cells. So what are our real goals here?
 * March 5, 2008

Shielding idea versus transport -	patent already held: use of melanin nanospheres o	how would the shielding take place, who/ with what structure o	how could the melanin be produced o	where would the suits be used? Where do they fit into society? o	New technology – can we find enough data for technical details and development o	Organic + synthetic : how to interface between the two
 * March 13, 2008 - Day After 3 Ideas Presentation

Aquaporins photosynthesis combination w/ photovoltaics -	osmosis would not be an efficient force to rely upon for powering the filter. (but better than gravity) -	Electric current applied to water, what would it do?

Pond w/ aquaporin scum -	large scale vs. small scale applications, which allows more freedom to design? -	Compare cost efficiency for this method of powering water purification, what level of efficiency, etc.

Group decision: Go with the combination idea for water filtration powered by photovoltaic cells. How will the current drive the water pump? Will E-field draw water molecules through? Water is full of ions - this could be used in an electric field. Other option is mechanical force from photovoltaic cells. To look up: microfluidics channels "concentrator" - how to filter the proteins out. What do we want to filter out? http://www.lsbu.ac.uk/water/magnetic.html

“Organic photovoltaics are flexible, lightweight, and potentially less expensive than traditional solar cells (they're "organic" because they're based on carbon). The main drawback is that organic PV cells are nowhere near as efficient at converting light into electricity as silicon cells. A recent development at Georgia Institute of Technology, however, is starting to close that performance gap. By adding a chemical called pentacene to the carbon "buckyballs" (Fullerenes again!) used in making the organic solar cells, the researchers were able to boost the efficiency to nearly 3.4 percent, with signs they could get to 5 percent in the near future. This compares to 25 percent for silicon cells (and up to 50 percent for experimental materials).”
 * SPRING BREAK RESEARCH:

“Although organic solar cells aren't as efficient, their other characteristics -- flexibility, weight, ruggedness, cost -- still make them attractive. They can be more readily embedded in other materials, from fabrics to plastics to roofing, and are ideal for small, low-power projects such as remote sensors. If, a decade from now, you drive a car with solar cells on its roof to help recharge the hybrid batteries, these are mostly likely the cells you'll be using.” From www.worldchanging.com/archives/001733.html

Ordered Nanostructures for Organic Photovoltaic Cells - www.nnin.org/doc/2004NNINreuMiller.pdf

Highlights of How OPvCs Work - http://kottan-labs.bgsu.edu/Presentations/pres23d.htm

News Article on Company combining opvcs into household products. http://www.news.com/newsblog/?keyword=Konarka

Average human consumes 1.5 gallons of water per day

Solar powered mechanical pump: cost = $2,500 http://www.simplepump.com/motorized_pumps.html

A water pump for the third world, heat-powered. http://www.thenakedscientists.com/HTML/content/interviews/interview/501/

Cheaper solar mechanical pumps: $1,500 http://store.solar-electric.com/cosofopipu.html

Step-by-step view from a water molecule’s point of view: 1.	pond 2.	dirty water basin 3.	through aquaporin in root system 4.	up through root by photovoltaic powered pump 5.	into clean water basin
 * April 1, 2008:

Questions: Should we dump the water through the root system rather than relying on diffusion, some sort of flow? For a small community, ‘water tower’ type system is good. Pump to assist flow is fine. Genetic question: how to create the aquaporin root system? Tank maintenance. More specific genetic questions: Aquaporin embedding + aquaporin manufacture/production

To be researched: -Polymer to embed aquaporin proteins in, we want something that will block ‘everything else’ from getting through

System breakdown: -Special cell membrane -light → power -light, resources, N + C -aquaporin to h2o
 * April 2, 2008:

DEVICES: Necessary: Aquaporins Way for cells to attach to each other w/o dirty water passing between, i.e. tight junctions? Possible: Ion pump? Cell pump to pump out cleaned h2o (active transport)? Ion regulator?

Concerns about cells: Ion concentration within cell – how can we make sure that the cell continues to pump thru aquaporins on one side considering that ion concentration will be higher on one side of the cell. (hence, possible need for ion regulator?) How to achieve what we want without “overloading” the cells, i.e. disrupting their normal balance/ internal processes. We need to supply nutrients, light. Kill mechanism to make sure cell growth doesn’t get out of our control, i.e. pesticides to kill or chemicals of some kind? How “start” the cells on the right track w/ our filtering process? Keeping a membrane-like structure – Kiran says: “should probably take care of itself?” Two Articles on Active Water Pump Protein in Plant Cells (Maize - corn) http://www.blackwell-synergy.com/doi/abs/10.1111/j.1399-3054.1966.tb07067.x http://jxb.oxfordjournals.org/cgi/reprint/24/1/33

Device List: 1.	aquaporins 2.	adhesor 3.	active transport of water protein 4.	ion pump

Tasks: Aquaporin – which one? PIP2? Ion pump – which one? Several? Danger of cell over-saturation/overload? Start/stop 	switch? Adhesor – which one? How thick/ how much? Active water transport – which one?

Research assignments: Kiran: adhesor and active ion pump, pp slides – go/no go, impact Benji: aquaporin, pp slides - debugging/testing, $, time Paul: active water pump, pp slides – issues, buildable


 * April 15, 2008:

Today, we made a group decision to focus on the aspect of creating a filtering membrane structure, using tight junctions, which will allow for creation of desired concentration gradient.

-Natural mechanism of attachment of cyanobacteria -Other ways for attachment -3 types: occluding communicating: allow ion ‘communication’, transfer anchoring: of cytoskeleton between cells Holdfasts: keep cyanobacteria in rods “Occluding junction-making protein”, claudins, others -Sheets, use communicating tight junctions between layers occluding tight junctions between cells within same layer Crux of our project: LOCALIZATION of Transmembrane Proteins

Cytoplasmic regulation of tight-junction permeability: effect of plant cytokinins http://ajpcell.physiology.org/cgi/content/abstract/239/3/C75

Arabidopsis group ‘Ie formins’ localize to speciﬁc cell membrane domains, interact with actin-binding proteins and cause defects in cell expansion upon aberrant expression http://www.blackwell-synergy.com/doi/pdf/10.1111/j.1469-8137.2005.01582.x

Distribution of Cell Membrane-associated Proteins Along the Human Nephron http://www.jhc.org/cgi/content/full/46/12/1423

The focus today was on scheduling and making a very detailed breakdown of what needs to get done over the next few weeks in terms of research, decision making, and write ups. The day-by-day breakdown calendar has been uploaded separately.
 * April 16, 2008:

TO DO LIST: -Decide what model organism to use - Green Algae or Spirulina? -Gap junctions: what can flow between cells? -Tight junction -Making sure that the algae will form sufficient number of junctions between cells to be impermeable. -How many layers do we want? What max/min level will be functions/sturdy enough? -Troubleshooting: will adding these junctions interfere w/ normal cell function? (We should at least have ways to test this.) Also, ratio of gap junctions and tight junctions: how many of each do we want/can we handle? -(Devices: feedback loops/mechanisms for control wrt to ion concentration inside cells.) -Finding the sequences -Presentation organization

Structure of membrane: Go with the ‘block’ idea because we don’t need to deal with cell division issues. Light needs to get through to the cells on the inside of the block, find out the limiting amount of stacking. Decision: use spirulina as host organism What will kill our cells and wreck the system? What is the tensile strength of the junctions between cells? i.e. How strong is our membrane physically? What if plasmodesmata do not form?
 * April 17, 2008:

Optimum Growth Conditions and light utilization efficiency of Spirulina platensis http://www.springerlink.com/content/n5g61618x8178887/

Would there be any other use for cell wall besides structure? Will removal harm cell processes? We would like to use tight junctions and gap junctions for our membrane. However, we also want the sheet to be photosynthetic so that it will be easy to maintain. But we don't want the cell walls to interfere with our gap and tight junctions. Can we induce L-form photosynthetic bacteria that would be viable for our sheet? Knock out cellulose synthase genes to prevent formation of cell wall. Is cellulose needed elsewhere in cell. From today's research, it seems no.
 * April 23, 2008:

From Cellulose to Cell - potentially good diagrams for our final presentation http://jeb.biologists.org/cgi/reprint/202/23/3263/b.pdf

1.	Prevent cell wall formation via knockout of cellulose synthase genes. 2.	Program cell for tight junctions and gap junctions.

Genes involved in tight junction + gap junction formation were found on NCBI database. Question: will these genes be directly transferable to our bacteria? i.e. actually produce the junctions?

Decided on organism: Synechococcus elongates
 * April 24, 2008:

Lab research questions/tasks: 1.	Assemble the gene sequence with promoter, genes, terminator, etc. 2.	Transform our host cells. 3.	Need a setup that resembles the environment of the real-world filtration setup – test ability of cells to grow on the polymer. 4.	Polymer/ Scaffolds – water-permeable. 5.	Tests for toxins/ what waste products do the cells excrete? 6.	How many aquaporins, ion pumps, tight junctions can the cell handle?
 * April 29, 2008:

This is the narrative summary of the project that I sent to Missy Holbrook the week before our phone interview.
 * April 30, 2008:

Basic Overview •	We want to purify water. We are planning on making a filter by producing sheets of photosynthetic cells (possibly some sort of plant cells). We would like our filter to be photosynthetic so that our filter is more self-sustaining and environmentally friendly. •	We want to move the water through the filter via aquaporins - through the cells - not around the cells through some sort of porous/size-selection process. We want to use aquaporins to move the water through the filter since aquaporins are perfectly selective for water and will allow nothing else to get through to the other side of the sheet. •	In order to make an effective filter of this sort, we must make our sheet impermeable to everything. •	We do not want to rely on a huge amount of pressure to push our water through the filter, so the water will move through the sheet with some sort of osmotic mechanism with an active water transport pump to move the water through the cells via the aquaporins. •	The osmotic gradient will be established by a system of ion pumps, channels, and regulators. The pumps will pump ions in from the dirty water into our filter cells to create a higher concentration of ions/other molecules compared to the dirty water outside. This will cause the water from the dirty water to flow into our cells. We must strictly control the concentration of ions present in the cell as to make it safe for the cell - controlled by an ion regulator of some sort enabling the opening and closing of ion channels.

Goal of the Current project •	Our project is only focused on producing the impermeable sheets/clumps of cells - the basic structure of the filter. We are planning to incorporate many components like aquaporins, an active water transport pump (which so far has not come up in our research), and ion pumps, but we are only designing how the cells will link to make the structure impermeable.

These are our questions, some of which we want to pose to Missy Holbrook: We would like to use tight junctions and gap junctions for our membrane. However, we also want the sheet to be photosynthetic so that it will be easy to maintain. But we don’t want the cell walls to interfere with our gap and tight junctions. Can we induce L-form photosynthetic bacteria that would be viable for our sheet?
 * Will removal of cell wall harm cell processes?
 * Genes involved in tight junction + gap junction formation were found on NCBI database. Question: will these genes be directly transferable to our bacteria i.e. actually produce the junctions?
 * Protein half-lives, does this mean we have to regulate?
 * What promoter should we use?
 * What is necessary for tight junctions? Claudins, occludins, JAMs.  TJPs and claudins: are these the only genes we need to insert into our bacteria?
 * Genes from what species would be best to insert our bacteria?
 * What are the “hypothetical proteins” that come up in NCBI gene searches?
 * How should we go about testing the setup in the lab – i.e. How many aquaporins, ion pumps, tight junctions can the cell handle?

Root Structure, Kill control device, inducers (explain purpose)
 * May 1, 2008:

Overall Paper Structure: 1.	Intro: what we’re doing and why. 2.	System description (diffusion and filter structure) – Here are the two aspects and why we need them. 3.	Components of system for each aspect of level two. 4.	Genetic makeup of each component and expression in cell. 5.	Specific recap of level two.

Six-month Plan:  Group A, Group B, Group C Aquaporin manipulation: evaluate each available aquaporin for best effiency. Cell wall manipulation: can cell wall production be localized to where we want it? -check for protein expression using fluorescence -use stain to check for desired localization Tight junctions: check for protein expression using florescence and verify junction locations using freeze-fracture and microscopy Find the optimal ratio of aquaporins to water pumps. Test via observation of cell shrinkage/expansion with varying numbers of each. Ensure pumps do not use too much energy – test different levels to see when cells die. Gap junctions: follow the same procedure as tight junctions Macro-testing: use modern water purity testing to make sure we filter out harmful compounds. All groups will test longevity of system and system maintenance i.e. that protein degradation rates equal protein synthesis rates, observe cell populations over time, observe filtration efficiency.
 * May 6, 2008

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