Team:NYU/Project

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Our game plan for the international genetically engineered machine competition is to construct a yeast strain capable of independent antibody discovery. Instead of relying on antibody display that require high-throughput fluorescent screening, currently the dominant method for microbial antibody discovery, our strain will be able to select for high antibody binding without outside influence. We will accomplish this by linking the antibody library and antigen with the split ubiquitin system, which will allow the yeast cells to sense the amount of antibody::antigen complexing.
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Our game plan for this year's competition is to construct an easy-to-use yeast strain capable of intracellular antibody discovery. The current dominant mode of antibody discovery using microbes requires cellular surface display and high-throughput fluorescent screening. Instead of using this method, we wanted our cells to be able to sense when the antibody they are translating will bind the target antigen. Using a modified form of the yeast two-hybrid screening system, the yeast cells will sense the amount of antibody::antigen interaction and will, in essence, screen themselves.  
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Basically, the single chain variable fragment (scFv) antibody will be fused to the N-terminal domain of ubiquitin (N-ub) and the target antigen will be fused to the C-terminal domain (C-ub). When the antibody and antigen form a complex, the two domains of ubiquitin are brought together and any protein that is fused downstream of C-ub will be cleaved from the rest of the complex by a ubiquitin protease. To use this mechanism to our advantage, we will fuse the Gal4 activator protein downstream of antigen::C-ub complex. So, when the antibody binds the antigen, Gal4p will be released, translocated into the nucleus and will affect transcription of genes that confer greater cell survival in the environment (either amino acid biosynthesis or, for a control, antibiotic resistance).
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In a research setting, once you have discovered the antibody you wish to use you must then reclone the coding region into a secretion vector for production of pure antibody protein. To make this process faster and more easily accomplished, we are also building a recombination-based architecture into our system that will allow the cells to go straight from screening to protein production in a matter of minutes. This concept has the capability to not only shave weeks off of current antibody discovery protocols, but opens the door to programmed cellular restructuring of antibody genes for different purposes.
== The Experiments ==
== The Experiments ==

Revision as of 20:42, 18 August 2010


Home Team Project BioBricks Modeling Notebook Safety


Russ, this should be the part where you talk about how you came up with the project idea.
NYU logo.png
This part should serve as the official abstract describing the project. Unless the two paragraphs in the project details section below is the abstract?
File:NYU team.png
Your team picture



Project Details

Our game plan for this year's competition is to construct an easy-to-use yeast strain capable of intracellular antibody discovery. The current dominant mode of antibody discovery using microbes requires cellular surface display and high-throughput fluorescent screening. Instead of using this method, we wanted our cells to be able to sense when the antibody they are translating will bind the target antigen. Using a modified form of the yeast two-hybrid screening system, the yeast cells will sense the amount of antibody::antigen interaction and will, in essence, screen themselves.

In a research setting, once you have discovered the antibody you wish to use you must then reclone the coding region into a secretion vector for production of pure antibody protein. To make this process faster and more easily accomplished, we are also building a recombination-based architecture into our system that will allow the cells to go straight from screening to protein production in a matter of minutes. This concept has the capability to not only shave weeks off of current antibody discovery protocols, but opens the door to programmed cellular restructuring of antibody genes for different purposes.

The Experiments

Results

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