What is it about?
We have created a new way to solve some of the mysteries among an increasingly important class of proteins that don’t appear to have any specific structures but serve very important functions, including the complex genetic processes that separate high-order organisms from single-cell bacteria. The method, which we call "Molecular LEGO", consists of pulling the proteins apart and rebuilding them, segment by segment to study how the parts interact with each other by both experiments and molecular simulations.
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Why is it important?
For the past four decades, biochemists and biomolecular engineers have used techniques such as X-ray crystallography, NMR and cryo-electron microscopy to study proteins that fold into defined structures that dictate how they work like the nanoscale machines they are. Scientists have also traditionally used a method to help understand what makes these proteins behave as they do that consists of creating mutations in the protein, changing single amino acids, and studying how much that change affects the protein’s structure, stability, and the rates of folding and unfolding. However, intrinsically disordered proteins (IDPs), discovered in the past 20 years, don’t have apparent structures — at least none that current techniques can easily discern. IDPs change shapes based on their environments and conditions and tend to fold into structures when they bind specific partner molecules. They have the unique ability to morph in response to multiple partners and can process sophisticated inputs and outputs. But it has been a mystery whether their response is passive — entirely determined by the partner — or controlled via an internal folding mechanism that has yet to be revealed. That is the question we are trying to address in this work by using a well-studied partially disordered protein called NCBD to uncover clues as to how it performs its sophisticated biological function as proof-of-concept. So we took this disordered protein and made it even more disordered by breaking it into pieces and studying each one separately. Then we recombined the segments in order, one at a time, to see how each restored, larger segment behaves. What we found is that, while ordered proteins behave as if they have on and off switches, IDPs seem to work more like rheostats, changing gradually.
Perspectives
It could turn out that IDPs only appear disordered because we are looking at them using techniques that don’t give us the whole picture. The IDPs must have some structure because they are able to select specific partners, change shape when bound to those partners and complete complicated actions in very specific ways. More of these IDPs are being discovered and are quickly becoming a very important class of proteins. They are more commonly found in high-order organisms, such as humans. It seems like the paradigm that is emerging is that all these proteins are key in regulation and responsible for all the complexity that is emerging in high-order organisms without having to vastly increase the number of genes. For example, an E. coli bacterium has about 5,000 genes, while a human has about 30,000. So, you can see that we definitely have to do something special with those 30,000 genes to make us, compared with a bacterium. The thought is that this is achieved by sophisticated regulation, networks and other complicated processes, and it seems like the key players in all this are these IDPs. They are often found at the hubs in these networks. This approach that we have developed could be instrumental to study them at new levels of detail. The next steps will be to apply the new technique to other proteins and to recombine the proteins outside the segment order defined by the gene sequence to see how that affects the segments and function. This connects with a lot of our engineering work in which we’re trying to build biosensors and new methods for diagnostics. We can use these proteins as the scaffolds to make responsive systems on the molecular level.
Victor Muñoz
University of California Merced
Read the Original
This page is a summary of: A modular approach to map out the conformational landscapes of unbound intrinsically disordered proteins, Proceedings of the National Academy of Sciences, June 2022, Proceedings of the National Academy of Sciences,
DOI: 10.1073/pnas.2113572119.
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