Team:ATOMS-Turkiye/Project/Module1/Goals

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==Goals & Challenges==
==Goals & Challenges==
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'''Module 1 of our project consists of four stages'''
 
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·        To detect cancer cells specifically using the nanofactory complex.
 
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Firstly, we had to devise a system which would detect the cancer cells. This challenge was solved by searching for a specific marker present on cancer cells; this was EpCaM(1). EpCAM markers are present on both healthy cells and cancer cells. However according to an article the number o EpCAM present on cancer cells are 100 times more compared to a healthy cell.
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''1• To detect cancer cells specifically using the nanofactory complex.''
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Nanofactories were the complexes we aimed to detect cancer cells. This complex consists of an Anti-EpCAM antibody that would bind to EpCAM antigens present on the cancer cells.We designed our bacteria as an effective alternative to presenting  antibody parts of our nanofactory on its surface. It was an obligation for us to use anti-EpCAM for binding. While we were researching on EpCAM, we found anti-EpCAM’S active domain called C215. C215 does the same job as Anti-Epcam though it is smaller than anti-EpCAM. Therefore, our choice was C215.
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With presenting C215 on the outer membrane, we designed a bacteria called “nanobacterium”. This bacteria will find the cancer cells and accumulate around them, which will call our bacteria towards the cancer cells.
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·        Producing AI-2 by our nanofactory without cease
+
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After detection we need a calling system to call the bacteria toward the cancer cells and we used a quorum sensing material called Auto Inducer-2(AI-2). AI-2 produced by our nanofactories call our bacteria towards the cancer cells which move via the taxis method.
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Firstly, we had to devise a system which would be able to detect the cancer cells. This challenge was solved by searching for a specific marker present on cancer cells and that was EpCaM[1]. EpCAM markers are present on both healthy cells and cancer cells. However according to an article[1] the number of EpCAM present on cancer cells are 100 times more compared to a healthy cell. Nanofactories were the complexes we aimed to detect cancer cells. This complex consists of an Anti-EpCAM antibody that would bind to EpCAM antigens present on the cancer cells.We designed our bacteria as an effective alternative to presenting antibody parts of our nanofactory on its surface. It was an obligation for us to use anti-EpCAM for binding. While we were researching on EpCAM, we found anti-EpCAM’S active domain called C215. C215 does the same job as Anti-Epcam though it is smaller than anti-EpCAM. Therefore, our choice was C215. With presenting C215 on the outer membrane, we designed a bacteria called “nanobacterium”. This bacteria will find the cancer cells and accumulate around them, which will call our bacteria towards the cancer cells.
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·        Expression of cancer specific apoptotic protein by the bacteria because of AI-2 concentration
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''2• Producing AI-2 by our nanofactory without cease''
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We need to trigger the cancer killer protein expressioning mechanism with a stimulant. This was a serious problem. We solved this challenge by using intentive AI-2 concentration near by of cancer cells.  The bacteria will sytnthesize our protein when AI-2 concentration reaches to a certain level. When the bacteria arrive to around the intensive concentration of AI-2, this situation triggers a mechanism of the bacteria which produces our killer protein.
+
After detection a system was required to call the bacteria towards the cancer cells, therefore we used a quorum sensing material called Auto Inducer-2(AI-2). AI-2 was produced by our nanofactories which enabled our bacteria to move towards the cancer cells via taxis method.
-
+
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·        Secreting the killer protein out of the cell with signal peptides.
+
''3• Expression of cancer specific apoptotic protein by the bacteria due to AI-2 concentration''
-
We used HLYA signal peptide to secrete our apoptotic proteins from the bacteria. We also wished to use TorA signal peptide as an alternative to HLYA (3). While we were using TorA, it was secreting our proteins to periplasm,not to extra cellular matrix. To solute this problem we used a chemical substance for pokes holes on the outer membrane. And our proteins released after this step.
+
We had to trigger the expression of our cancer killer protein with a stimulant. This complicated challenge was solved by using intentive AI-2 concentration near by of cancer cells. The bacteria will sytnthesize our protein when AI-2 concentration reaches to a certain level. Once the bacteria enter the environment of intensive concentration of AI-2, a mechanism is triggered which enable our bacteria to produce the killer protein.
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1)EpCAM overexpression prolongs proliferative capacity of primary human breast epithelial cells and supports hyperplastic growth
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Agnieszka Martowicz1, Johannes Rainer2, Julien Lelong2, Gilbert Spizzo1, Guenther Gastl13 and Gerold Untergasser3*
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''4• Secreting the killer protein out of the cell with signal peptides.''
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(2) Biological Nanofactories Target and Activate Epithelial Cell Surfaces for Modulating Bacterial Quorum Sensing and Interspecies Signaling
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Colin G. Hebert,†, Apoorv Gupta,§ Rohan Fernandes,†, Chen-Yu Tsao,†, James J. Valdes,‡ and William E. Bentley†,§
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We used HLYA signal peptide to secrete our apoptotic proteins from the bacteria. We also wished to use TorA signal peptide as an alternative to HLYA [3]. While using TorA, our proteins were secreted to the periplasm and not to the extra cellular matrix. As a solution to this problem we used a chemical substance which pokes holes on the outer membrane proteins and release our proteins inside.
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3.) Improved secretory production of recombinant proteins by random mutagenesis of hlyB, an alpha-hemolysin transporter from Escherichia coli.
+
 
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(1)EpCAM overexpression prolongs proliferative capacity of primary human breast epithelial cells and supports hyperplastic growth Agnieszka Martowicz1, Johannes Rainer2, Julien Lelong2, Gilbert Spizzo1, Guenther Gastl13 and Gerold Untergasser3*
 +
(2) Biological Nanofactories Target and Activate Epithelial Cell Surfaces for Modulating Bacterial Quorum Sensing and Interspecies Signaling Colin G. Hebert,†, Apoorv Gupta,§ Rohan Fernandes,†, Chen-Yu Tsao,†, James J. Valdes,‡ and William E. Bentley†,§
 +
(3) Improved secretory production of recombinant proteins by random mutagenesis of hlyB, an alpha-hemolysin transporter from Escherichia coli.

Latest revision as of 12:46, 20 May 2014

Goals & Challenges

1• To detect cancer cells specifically using the nanofactory complex.

Firstly, we had to devise a system which would be able to detect the cancer cells. This challenge was solved by searching for a specific marker present on cancer cells and that was EpCaM[1]. EpCAM markers are present on both healthy cells and cancer cells. However according to an article[1] the number of EpCAM present on cancer cells are 100 times more compared to a healthy cell. Nanofactories were the complexes we aimed to detect cancer cells. This complex consists of an Anti-EpCAM antibody that would bind to EpCAM antigens present on the cancer cells.We designed our bacteria as an effective alternative to presenting antibody parts of our nanofactory on its surface. It was an obligation for us to use anti-EpCAM for binding. While we were researching on EpCAM, we found anti-EpCAM’S active domain called C215. C215 does the same job as Anti-Epcam though it is smaller than anti-EpCAM. Therefore, our choice was C215. With presenting C215 on the outer membrane, we designed a bacteria called “nanobacterium”. This bacteria will find the cancer cells and accumulate around them, which will call our bacteria towards the cancer cells.

2• Producing AI-2 by our nanofactory without cease

After detection a system was required to call the bacteria towards the cancer cells, therefore we used a quorum sensing material called Auto Inducer-2(AI-2). AI-2 was produced by our nanofactories which enabled our bacteria to move towards the cancer cells via taxis method.

3• Expression of cancer specific apoptotic protein by the bacteria due to AI-2 concentration

We had to trigger the expression of our cancer killer protein with a stimulant. This complicated challenge was solved by using intentive AI-2 concentration near by of cancer cells. The bacteria will sytnthesize our protein when AI-2 concentration reaches to a certain level. Once the bacteria enter the environment of intensive concentration of AI-2, a mechanism is triggered which enable our bacteria to produce the killer protein.

4• Secreting the killer protein out of the cell with signal peptides.

We used HLYA signal peptide to secrete our apoptotic proteins from the bacteria. We also wished to use TorA signal peptide as an alternative to HLYA [3]. While using TorA, our proteins were secreted to the periplasm and not to the extra cellular matrix. As a solution to this problem we used a chemical substance which pokes holes on the outer membrane proteins and release our proteins inside.

(1)EpCAM overexpression prolongs proliferative capacity of primary human breast epithelial cells and supports hyperplastic growth Agnieszka Martowicz1, Johannes Rainer2, Julien Lelong2, Gilbert Spizzo1, Guenther Gastl13 and Gerold Untergasser3* (2) Biological Nanofactories Target and Activate Epithelial Cell Surfaces for Modulating Bacterial Quorum Sensing and Interspecies Signaling Colin G. Hebert,†, Apoorv Gupta,§ Rohan Fernandes,†, Chen-Yu Tsao,†, James J. Valdes,‡ and William E. Bentley†,§

(3) Improved secretory production of recombinant proteins by random mutagenesis of hlyB, an alpha-hemolysin transporter from Escherichia coli.