Antibodies

Antibodies are proteins produced by the immune system of vertebrates as a response to harmful substances. They recognize and bind to antigens found in bacteria, viruses, fungi, or toxins, thereby helping to protect our body. Beyond health, antibodies are indispensable research tools, used in techniques like Western Blots, ELISAs and flow cytometry, as well as in medicine through vaccines and therapeutic drugs.

Variable Lymphocyte Receptors

Nature already offers an answer. Jawless fish, including lampreys, like our mascot Larry the Lamprey, do not use antibodies at all. Instead, their immune system relies on Variable Lymphocyte Receptors (VLRs).
VLRs function like antibodies, but they are built from repeating structural modules called variable Leucine-Rich Repeats (LRRVs). These repeats determine the binding behavior of the protein through changing the structure of the concave face of VLRs, mediating immune response.

However, VLRs are complex and difficult to express in bacteria, which poses challenges for scalable production. To overcome this problem, a small change in structure is necessary.

Repebodies

Repebodies are modified versions of VLRs and specifically designed to function in E. coli. Replacing their N-terminal cap with that of the bacterial protein Internalin-B improves their solubility and production efficiency.
This breakthrough means that Repebodies can be easily produced in bacteria, unlike antibodies. Now that we have a scalable alternative to antibodies, the question is...

Creating Diversity

To explore the huge potential of Repebodies, we need diversity. Our approach introduces mutations directly into the genes that code for Repebodies, thereby creating a vast library of variants.

Introducing Mutations

• Short single-stranded DNA (ssDNA) fragments carrying the desired mutations are introduced into the cell.
• During DNA replication, these ssDNAs anneal to the lagging strand, creating mismatches that lead to mutations in the leucine-rich repeat regions of the Repebodies.
• Over time and multiple cell divisions, these mismatches are permanently incorporated, generating a diverse set of Repebody variants.

Retrons

Retrons are natural elements that produce ssDNAs directly inside the cell. This allows our entire diversification system to work fully in vivo, without external DNA handling.

This process creates a large pool of different Repebodies, each with unique binding capabilities.

Selection

Diversity is only useful if we can find the proteins we actually want. That is why we developed a high-throughput selection pipeline capable of screening large cultures to identify Repebodies that specifically bind to any protein of interest.
To optimize this process, we tested different readout systems for selection, including the bacterial adenylate cyclase two-hybrid (BACTH) system.

Learn more about our selection platforms

Targeted Protein Degradation

Targeted Protein Degradation (TPD) involves hijacking the cell’s natural degradation machinery to selectively eliminate a protein of interest (POI). This is a recent alternative to protein inhibition.
The great advantage: TPD aims to degrade the protein entirely, instead of just inhibiting its functions. The process involves designing a targeting chimera, which can bind both the POI and a degradation initiator (DI), pulling the target into the degradation machinery.

The POI ligands often consisted of:

• nanobodies, which requires antibodies
• synthetic parts, which cannot be expressed in E. coli, essentially requiring a delivery system

Repebodies in TPD

Repebodies, our alternative to antibodies, can be easily expressed in E. coli and tailored to a protein of interest using our mutagenesis approach. Integrating Repebodies as the POI ligand avoids the production of synthetic parts or the need for ordering a nanobody specific to your POI.

This way you can degrade your protein of choice with our Repebody!

Our project CONCAVE

We offer a toolkit for the easy creation of Repebodies with desired binding specificity to be used as an antibody alternative, combined with an application in targeted protein degradation.
This includes:

• Specific strain of E. coli with an integrated Repebody sequence optimized for randomization
• Plasmids encoding for the retron and selection system
• A template degrader construct for Targeted Protein Degradation

Our Toolkit

A powerful research tool for studying protein interactions and cellular processes Increase accessibility of advanced analytics, such as assays and diagnostics to detect and quantify target proteins Aid in the creation of new and innovative therapeutics based on targeted protein degradation or modulations

With our Repebody Toolkit, you can create your own high-affinity binders in E. coli, anytime, anywhere!