Putting MP3 in your cells

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Today, a blast from the past, in the form of an MP3.

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Putting MP3 in your cells

Do you remember when you would upload songs in MP3 format to your iPod to listen to them? Today’s paper is completely unrelated, but I can't help wondering if the authors had a second thought about the name they chose. Let’s get to it!

Practically all biological processes rely on protein-protein interactions (PPIs), whether it’s cell signaling, metabolism, immune response, gene expression, pretty much everything. In nature, the variety and complexity of these systems is staggering, but when it comes to synthetic biology, our protein circuits are still pretty basic. Why? Mainly because the domains we use to create these interactions are few and often prone to “crosstalk”: unwanted, non-specific interactions. To build more sophisticated protein network, we need large-scale libraries of orthogonal protein interaction domains ones that don’t interfere with each other. Researchers have made progress by developing coiled-coil heterodimers (proteins that interact through complementary sequences), but creating large sets is still challenging.

As you might have guessed, today’s paper set to solve this problem. The team here introduced MP3-seq, a high-throughput yeast two-hybrid (Y2H) method that uses sequencing to measure PPIs on a massive scale. MP3-seq can measure over 100,000 PPIs in a single experiment, by encoding protein pairs with DNA barcodes, and then using the barcode enrichment as a proxy measure for interaction strength.

In short, in this system protein pairs are fused are fused with a DNA-binding domain or an activation domain, and their interaction triggers the expression of a selectable marker (His3), which promotes yeast growth. By sequencing the DNA barcodes before and after selection provides an enrichment score, which reflects the strength of the protein interaction.

The team put MP3-seq to the test by validating it against well-characterized orthogonal coiled-coil interactions and human Bcl-2 proteins with de-novo designed inhibitors. MP3-seq showed a great ability to measure these interactions, correlating well with biochemical data.

But they didn’t stop here: the researches screened a huge library of designed heterodimers, multi-helical protein complexes, measuring over 100,000 PPIs. Pretty crazy! This large-scale screening identified both on-target interactions (the ones they were aiming for) and off-target ones (unexpected pairings). Using these data, the team explored design rules to create orthogonal protein interactions, where proteins bind only to their designated partner, and not other proteins. They optimized structural features like helical length and hydrogen-bond networks, making the interactions even more specific.

Finally (this is a juicy paper) the researchers also put AlphaFold and Rosetta (computational tools for predicting protein structures) to the test on this massive dataset. While these models gave valuable insights into protein interactions, they still couldn’t fully predict orthogonality or the effects of mutations—this is something that will still need to be experimentally tested.

All in all, this is a very cool paper, and extremely dense. High-throughput systems are becoming more and more important, especially in this age of increasing data hunger. There are a huge number of fields where systems like these can be helpful, for example:

• Drug discovery: MP3-seq could help identify new drug targets by screening PPIs involved in disease pathways.

• Synthetic biology: For engineered cells to behave predictably, we need reliable protein circuits. MP3-seq is a powerful tool for this.

• Gene therapy: PPIs are crucial in designing gene-editing tools like CRISPR/Cas9 and similar. MP3-seq could help refine these systems by mapping protein interactions, reducing off-target effects, and improving efficiency.

But these are just some of my thoughts about it! Go and read the article for yourself here!

In other news:

• Controlling fish crystals: Apparently, functions as different as vision and camouflage in zebrafish are based on biogenic crystals. This paper now shows how the shape of these crystals is genetically controlled by their chemical composition. Pretty cool stuff!

• Mimicking spiders: If your next Halloween costume is Spider-Man, rejoice! In this paper, researchers developed a new way to create artificial silk, by closely mimicking the natural spinning process of spiders. The resulting biomaterial has great properties for use in health applications. They also created artificial spiders, because why not have more of those?

• CRISPR/Cas Nanorobots: What is better than CRISPR/Cas? Combining it with gold nanoparticles to make nanorobots, of course. In this study, the team studied the trans cleavage activity of Cas12a and then created nanorobots that improve the cleavage kinetics, leading to successful real-time imaging of microRNA in live cells.

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