Freiburg

From 2007.igem.org

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Natalia Maier
Natalia Maier
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[https://2007.igem.org/User:Dario Hermida Aponte]
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[https://2007.igem.org/User:Dario Dario Hermida Aponte]
Philipp Mappes
Philipp Mappes
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Andreas Hiltbrunner, Michael Reth, Bodo Rak
Andreas Hiltbrunner, Michael Reth, Bodo Rak
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== Our Project: ==
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== Project Summary ==
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Our goal is to design integrated molecular sensing and executing devices based on modular protein engineering. These integrated devices can then easily be used for the construction of complex systems. We plan to fuse sensing proteins, which provide nano-mechanical movements upon an external signal, to executing proteins, which depend in their activity on the nano-mechanical movement of the sensing protein. To elucidate the possibilities of such a system we plan to fuse each of the termini of Calmodulin, which senses Ca2+, or the Maltodextrin Binding Protein, which senses maltodextrin, to one half of a split enzyme such as β-lactamase. We expect that we can design the separation of the two enzyme fragments by one conformation of the sensing protein such that there is no activity and that a conformational change in the sensor will bring the enzyme fragments together resulting in activity of the execution part. Once the basis of the interconnection of sensor and executor has been established, we plan to extend the work in a modular fashion to further proteins in order to generate a readily available and easy to tailor toolbox. We see application of such tools as varied as bacterial growth regulation and tumor targeting.
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<b>Integrated Sensor-Executor Proteins and Molecular Switches</b><br>
 +
Our goal is to design integrated molecular sensing and executing devices based on modular protein engineering. These integrated devices can then easily be used for the construction of complex systems. We fuse sensing proteins, which provide nano-mechanical movements or dimerization upon an external signal, to executing proteins, which depend in their activity on the nano-mechanical change in the sensing part.<br>
 +
To elucidate the possibilities of such a system we used the calcium-ion sensor Calmodulin and the light sensor system PhyA-Fhy1. To test execution we used the split enzymes DHFR or beta-lactamase or the fluorescent proteins CFP and YFP, which can form a FRET pair. Sensors and executors were geneticlly fused and tesetd in E. coli for activity. So far we could demonstrate Ca2+ dependent growth of E. coli with the DHFR[1]-Calmodulin-DHFR[2] construct.
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<br>
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[[Freiburg07/report_Ca_sensor| final iGEM report: Ca2+ sensor]]<BR>
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[[Freiburg07/report_light_sensor| final iGEM report: light sensor]]<BR>
 +
 
 +
 
 +
<b>Biobrick compatible strategy for fusion proteins</b><br>
 +
The present BioBrick prefix and suffix rules are not compatible with modular protein design. Thus, we propose an extension of the present standard for fusion proteins in which two restriction sites are added in frame adjacent to the coding sequence.<br>
 +
[[Freiburg07/report_fusion_parts| final iGEM report: Fusion parts]]<BR>
== Support: ==
== Support: ==
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[http://www.syntheticbiology.ethz.ch/synbiocomm/index SYNBIOCOMM]
 
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[http://www.uni-freiburg.de/ Universität Freiburg]
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[http://www.syntheticbiology.ethz.ch/synbiocomm/index SYNBIOCOMM]<br>
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[http://www.uni-freiburg.de/ Universität Freiburg]<br>
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[http://www.uni-freiburg.de/wiss-ges/ Wissenschaftliche Gesellschaft in Freiburg im Breisgau]<br>
 +
[http://www.geneart.com GeneArt]<br>
== Lab-Work: ==
== Lab-Work: ==
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[[Freiburg07/labnotes| lab notes]]<BR>
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[[Freiburg07/labnotes1| lab notes1]]<BR>
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[[Freiburg07/labnotes2| lab notes2]]<BR>
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[[Freiburg07/labnotes| lab notes3]]<BR>
[[Freiburg07/plasmids| plasmids]]<BR>
[[Freiburg07/plasmids| plasmids]]<BR>
[[Freiburg07/primers| PCR primers]]<BR>
[[Freiburg07/primers| PCR primers]]<BR>
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== Protocols: ==
== Protocols: ==
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[[Freiburg07/Mediums and Plates| Mediums and Plates]]<BR>
[[Freiburg07/DNA_sequencing| DNA sequencing]]<BR>
[[Freiburg07/DNA_sequencing| DNA sequencing]]<BR>
 +
[[Freiburg07/Ligation| Ligation]]<BR>
[[Freiburg07/Plasmidprep| Plasmid spin column prep]]<BR>
[[Freiburg07/Plasmidprep| Plasmid spin column prep]]<BR>
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[[Freiburg07/Glycerol_stock| Glycerol stocks]]
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[[Freiburg07/Glycerol_stock| Glycerol stocks]]<BR>
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[[Freiburg07/Gene_Protein_info| General Gene-Protein Information]]<BR>
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[[Freiburg07/Purification|Purification ]]<BR>
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[[Freiburg07/Transformation|Transformation ]]<BR>
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[[Freiburg07/Dephosphorylation|Dephosphorylation ]]<BR>
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[[Freiburg07/Preparative Digestion|Preparative Digestion ]]<BR>
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[[Freiburg07/Analytic Digestion|Analytic Digestion ]]<BR>
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[[Freiburg07/Gel Electrophoresis|Gel Electrophoresis ]]<BR>
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[[Freiburg07/Polyacrylamide gel electrophoresis|Polyacrylamide gel electrophoresis ]]<BR>
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[[Freiburg07/Protein purification|Protein purification ]]<BR>
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[[Freiburg07/In vivo test I|In vivo test I]]<BR>
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[[Freiburg07/In vivo test II|In vivo test II]]<BR>
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== Sandbox - wiki test: ==
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[[Freiburg07/sandbox| sandbox]]<BR>
<!-- Hi allerseits -falls jemand das vor Freitag liest- hätte gerne Feedback:
<!-- Hi allerseits -falls jemand das vor Freitag liest- hätte gerne Feedback:
-wie detailliert soll das projekt hier beschrieben werden? andre teams halten damit eher hinterm berg...?
-wie detailliert soll das projekt hier beschrieben werden? andre teams halten damit eher hinterm berg...?

Latest revision as of 14:55, 25 October 2007

BanneriGem.gif

Contents

Team Members:

Picture: The University of Freiburg iGEM Team in June 2007 (left to right: Natalia, Dinah, Corinna, Philipp, Katja, Maximilian, Moritz, Dario, Kristian)

Instructors:

Kristian M. Müller, Katja M.Arndt

Students:

Maximilian Mauler

Moritz Busacker

Corinna Gruber

Natalia Maier

Dario Hermida Aponte

Philipp Mappes

Graduate Student:

Dinah Mattay

Advisors:

Andreas Hiltbrunner, Michael Reth, Bodo Rak

Project Summary

Integrated Sensor-Executor Proteins and Molecular Switches
Our goal is to design integrated molecular sensing and executing devices based on modular protein engineering. These integrated devices can then easily be used for the construction of complex systems. We fuse sensing proteins, which provide nano-mechanical movements or dimerization upon an external signal, to executing proteins, which depend in their activity on the nano-mechanical change in the sensing part.
To elucidate the possibilities of such a system we used the calcium-ion sensor Calmodulin and the light sensor system PhyA-Fhy1. To test execution we used the split enzymes DHFR or beta-lactamase or the fluorescent proteins CFP and YFP, which can form a FRET pair. Sensors and executors were geneticlly fused and tesetd in E. coli for activity. So far we could demonstrate Ca2+ dependent growth of E. coli with the DHFR[1]-Calmodulin-DHFR[2] construct.
final iGEM report: Ca2+ sensor
final iGEM report: light sensor


Biobrick compatible strategy for fusion proteins
The present BioBrick prefix and suffix rules are not compatible with modular protein design. Thus, we propose an extension of the present standard for fusion proteins in which two restriction sites are added in frame adjacent to the coding sequence.
final iGEM report: Fusion parts

Support:

[http://www.syntheticbiology.ethz.ch/synbiocomm/index SYNBIOCOMM]
[http://www.uni-freiburg.de/ Universität Freiburg]
[http://www.uni-freiburg.de/wiss-ges/ Wissenschaftliche Gesellschaft in Freiburg im Breisgau]
[http://www.geneart.com GeneArt]

Lab-Work:

lab notes1
lab notes2
lab notes3
plasmids
PCR primers
Lab Boxes

Protocols:

Mediums and Plates
DNA sequencing
Ligation
Plasmid spin column prep
Glycerol stocks
General Gene-Protein Information
Purification
Transformation
Dephosphorylation
Preparative Digestion
Analytic Digestion
Gel Electrophoresis
Polyacrylamide gel electrophoresis
Protein purification
In vivo test I
In vivo test II

Sandbox - wiki test:

sandbox

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