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Targeted Protein DegradationUbiquitination Cells make, regulate and break down proteins constantly. They have a system to control the regulation of the proteins they produce. This is to remove unwanted proteins when no longer used. It also maintains healthy proteins as they degrade slowly over time. The process of ubiquitination is the tagging of these proteins by the cell for destruction. There are 3 enzymes that work in the process of ubiquitination.  They are the enzymes E1, E2 and E3. There are only 2 types of E1 enzymes, 30 types of E2 enzymes and estimated to be 600 possible E3 enzymes. Lets take a look at how a protein get tagged for destruction.  The E1 ligase take the actual ubiquitin molecule and puts a phosphate on it. This is called phosphorylation. Once its got that phosphate group added, it passes the ubiquitin to the E2 enzyme. The E2 ligase takes the phosphorylated ubiquitin and binds to the E3 ligase with it. This creates a complex of the E2 and E3 ligases. The E3 ligase will bind to the protein it recognizes and add that ubiquitin to the protein. The addition of the ubiquitin to the protein is done on a lysine amino acid. It can be done to several lysine amino acids on that same protein by multiple E3 ligases. After the E3 adds the ubiquitin to the protein, it releases the E2 ligase and binds another which contains another ubiquitin. It keeps adding these ubiquitin molecules to the lysine in a long chain called a poly ubiquitin chain. It can create a chain of more than 10 ubiquitin molecules long. This long tail of ubiquitin on the protein flags it for destruction by the organelle called the Proteasome. The Proteasome The Proteasome is a cellular organelle. Its like the recycling bin for proteins. When a cell is done with a protein, it tags it for destruction in the process called ubiquitination. The proteasome will load these tagged proteins and break them down into peptides of about 7 to 10 amino acids in length for recycling. They will further be broken down after into single amino acids for reuse to build new proteins.  The proteasome is made up of 3 subunits. The central 20s subunit is called the catalytic barrel. This is the business part of the proteasome where it actually breaks down the peptide chain of amino acids into smaller peptides.  The top an bottom of the 20s subunit has a 19s subunit bound to each end. These are designed to recognize the poly ubiquitin tails on proteins and load them into the barrel for shredding. The 19s subunit will also be responsible for unfolding the proteins into a single long peptide chain so that it feeds smoothly into the catalytic barrel. Together the 3 subunits make up the full 26s proteasome structure. The 3 subunits are not always assembled in the cell. They only come together when they are directed to do so. Protein Degraders There are 2 types of protein degraders in development and a 3rd in concept phase of development. The first is the monoDAC, the second is the biDAC and the last is the triDAC. The monoDAC will bind with a covalent chemical bonding to the E3 ligase and alter its targeted function. It changes the shape of the E3 and directs it to place the ubiquitin molecule onto a protein it directs. The monoDAC is typically smaller and has some advantages due to its size. It can also have more off target effects due to how it works. These are also referred to as mono valent or molecular glues.  The biDAC has 2 molecules that are held together by a linker. The first targets the desired E3 ligase and the other targets the desired protein. This bring together the right protein to the right E3 for ubiquitination. These are often called bi valent or PROTACs which stand for Proteolysis Targeting Chimera. They tend to be much larger than the monoDAC, but have more accuracy. The last one is mostly in concept, but its a trivalent protein degrader. It bind to the E3 ligase and has a linker to 2 different proteins.  Molecular Glue In previous Targeted Protein Degrader threads, I went over the basic process of how the E1 enzyme adds the phosphate group to the ubiquitin molecule. It then passes it to the E2 enzyme which binds to the E3 ligase as a complex.  The E3 ligase is designed with a site of recognition that is specific to a group of proteins. These proteins, called substrates, can be many different proteins for the same E3 ligase, but they all have a specific site that the E3 recognizes and binds to.  The molecular glue is a very small molecule that binds to a protein and provides a site to which the E3 ligase will bind. This allows for proteins that would never normally bind to that ligase to bind for targeted degradation.  This makes the molecular glue very small and powerful. The biggest drawback is that all of the current molecular glues that have been discovered have been by accident. This is one space where AI might dramatically improve this science. The biggest potential risk is they are so small, and they bind to a protein. This means there could be off target biding of proteins that resemble the target protein leading to off target side effects. |
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