Team:TU Dresden/Project/Description

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Description

Introduction

Our project is called SPACE-P (Structural Phage Assisted Continuous Evolution of Proteins) and is part of the Synthetic Biology field. It focuses on accelerating the process of developing protein binding partners, which is required in pharmaceutical research as well as biotechnology and many other fields of science. Thus far phage display is the most commonly used method for the discovery of protein binding partners. In this method a large library of potential binding partners is created where the strongest candidates are selected for further affinity testing. This process is very time consuming, cost intensive, requires human intervention steps, and is limited to the size of the given library. Our platform will make it possible to start with a single molecule, transform it by mutation and selection pressure to improve the binding affinity towards a target protein. This method is not only easy to implement, utilizing simple organisms and devices, but also significantly faster and cheaper than currently used tools.

Background

PACE

During the development of our project we had four meetings with the group leaders of the Biotechnology Center of the TU Dresden (BIOTEC). These meetings take place the first Monday of every month. The first one we attended focused on the idea we wanted to realize. The leaders helped us to think about difficulties we could face and gave valuable impulses. In the second one we presented our finance plan and they confirmed their support for an iGEM team. The third and fourth meeting were used to present the progress of our project as well as to check the finances. To sum up, we got input from diverse groups in the BIOTEC concerning both technical issues and ways to fund our group. This helped us to illuminate the project idea from numerous points of view.Phage assisted continuous evolution or PACE, is a system designed for the continuous, directed evolution of biomolecules. The principle is that E. coli flow through a lagoon in which M13 are present and viable. Important to note is that the flow rate of the E.Coli is adjusted so that it is faster than their doubling time but not of that of the M13. Another important aspect of this setup is the deletion of gene P3 on M13, which is necessary for infection and proliferation. Instead the gene P3 of the M13 is encoded on the E. coli plasmid, under the control of an upstream activating sequence (UAS). To undergo further infection cycles, the initial infectious generation of transgenic phage must activate the UAS by binding their activating domain (AD) to the binding domain (BD). This requires favorable protein-protein interactions between an X provided by the M13 and Y provided by the UAS of the E.Coli. As a result, and some additional help from a mutagenesis plasmid, M13 evolves this interaction between X and Yin order to stay in the lagoon. A stronger interaction creates a selection advantage and will be favored over a weaker interaction.