Intrinsically Disordered Proteins as Drug Discovery Targets

The intriguing world of the Intrinsically disordered proteins and why they are important as targets for drug discovery

When we think about proteins, we usually picture them as having a well-defined, three-dimensional, globular shape; however, this doesn’t apply to intrinsically disordered proteins or regions (IDPs/IDRs), which lack a fixed structure. IDPs/IDRs constitute at least 33% of the eukaryotic proteome.^1,6 IDPs are of significant interest to biotech and pharmaceutical companies because they are involved in many pathologies. As a result, IDPs have emerged as promising candidates for drug design.

In this blog we delve into the world of IDPs to understand their biophysics and their significance both to biology and drug design.

What are Intrinsically Disordered Proteins (IDPs)?

By definition, an IDP is a polypeptide chain that lacks a stable structure and remains highly dynamic under physiological conditions, a characteristic crucial for their biological functions. These proteins can adopt transient, non-globular conformations, possessing secondary or tertiary structures.^1,2,3 Short disorder regions are associated with flexible linker or loops, while long disordered regions are associated with recognition sites for protein – protein interactions due to their structural and functional independence from the rest of the protein core.¹

Where do IDPs exist in nature?

IDPs are found in the proteomes of all kingdoms of life and play vital roles in various biological processes, often by participating in protein-protein interactions that govern key cellular functions such as transcription and cell-cycle regulation. While all proteins are made of amino acids linked by peptide bonds, disordered proteins have a different amino acid composition compared to globular proteins, resulting in a wider range of conformational and structural heterogeneity. IDPs generally favor hydrophilic amino acids over hydrophobic residues. As a result, they are typically rich in arginine, glutamine, glutamic acid, lysine, proline, and serine, while being deficient in cysteine, isoleucine, leucine, phenylalanine, tryptophan, tyrosine, and valine.

The proportion of structural disorder increases with genome complexity, from bacteria to archaea to eukaryotes. Approximately 78% of eukaryotic proteins contain long intrinsically disordered regions (IDRs), compared to 12-14% in bacteria and 1% in archaea.

What processes do IDPs and IDRs participate in?

The primary roles of IDPs and IDRs are in signaling and regulation within and between cells. They are involved in processes such as post-translational modification, alternative splicing, cell division, nucleic acid replication and repair, and response to external stimuli. Their lack of stable structure allows IDPs to interact with multiple partners, facilitating complex signaling networks. Phosphorylation and other post-translational modifications (PTMs), which are predominantly located in IDRs, significantly influence their conformation and interactions^8,9. Additionally, alternative splicing often affects disordered regions, making it less disruptive compared to structured regions¹⁰.

Why IDPs Are Good Candidates for Drug Discovery?

IDPs play crucial roles in cellular functions, and their dysregulation can lead to diseases, particularly neurodegenerative disorders and cancer¹¹. The tendency of IDPs to form aggregates is a hallmark of numerous neurodegenerative diseases¹². For example, the IDP alpha-synuclein can form toxic aggregates that disrupt cellular functions, leading to Parkinson’s disease.

Targeting IDPs for therapeutics presents a series of challenges. Their natural dynamics make them difficult targets for traditional strategies, which typically involve small molecules binding to well-defined pockets.¹³ However, the ability of short amino acid sequences to bind diverse molecules suggests that rationally identifying small-molecule drugs targeting disordered proteins is feasible based on their sequence characteristics¹¹. Recent advances in drug discovery and protein engineering suggest that stabilizing certain IDP conformations could prevent protein-protein interactions and mitigate toxic aggregation, making IDPs potential therapeutic targets.

The structural plasticity of IDPs makes them difficult to study with traditional drug discovery biophysics approaches. However, Nuclear Magnetic Resonance (NMR) spectroscopy in solution is valuable for exploring protein-ligand interactions in IDPs for therapeutic purposes. These techniques are available within the Protein & Structure department at Sygnature Discovery.

Our experience in working with IDPs

 Understanding the structural and dynamic properties of proteins is crucial for drug discovery. Historically, most attention has been given to well-structured proteins. However, this subset of disordered proteins is now at the forefront of research.

Here at Sygnature Discovery, we pride ourselves on our extensive experience of working with these fascinating molecules. Our team possesses extensive expertise in protein biochemistry, cell science, structural biology, and biophysics, all dedicated to unraveling the complexities of IDPs and IDRs. We have led challenging projects related to IDPs, successfully expressing, purifying, and structurally characterizing these targets. Our successes include:

• Improvements in expression
• Improvements in solubility
• Degradation mitigation
• Improvements in yield
• Improvements in protein sample quality.
• Establishing SPR assays to measure compound binding
• NMR studies of the dynamics of IDPs

References

1. 1.- Trivedi R, Nagarajaram HA. Intrinsically Disordered Proteins: An Overview. Int J Mol Sci. 2022 Nov .
2. 2.- Uversky, Vladimir N. “Introduction to intrinsically disordered proteins (IDPs).” Chemical reviews
3. 3.- Arai, Munehito et al. “Dynamics and interactions of intrinsically disordered proteins.” Current opinion in structural biology vol. 84 (2024)
4. 4.- Uversky Vladimir N.Intrinsically Disordered Proteins and Their “Mysterious” (Meta)Physics.Frontiers in Physics, 7, 2019
5. 5.- Dunker AK, Babu MM, Barbar E, Blackledge M, Bondos SE, Dosztányi Z, Dyson HJ, Forman-Kay J, Fuxreiter M, Gsponer J, Han KH, Jones DT, Longhi S, Metallo SJ, Nishikawa K, Nussinov R, Obradovic Z, Pappu RV, Rost B, Selenko P, Subramaniam V, Sussman JL, Tompa P, Uversky VN. What’s in a name? Why these proteins are intrinsically disordered: Why these proteins are intrinsically disordered. Intrinsically Disord Proteins. 2013 Apr.
6. 6.- Basile W, Salvatore M, Bassot C, Elofsson A. Why do eukaryotic proteins contain more intrinsically disordered regions? PLoS Comput Biol. 2019 Jul 22.
7. 7.- Dunker, A.K., Cortese, M.S., Romero, P., Iakoucheva, L.M. and Uversky, V.N. (2005), Flexible nets. The FEBS Journal.
8. 8.- Wright, Peter E, and H Jane Dyson. “Intrinsically disordered proteins in cellular signalling and regulation.” Nature reviews. Molecular cell biology vol. 16,1 (2015).
9. 9.- Bondos, Sarah E et al. “Intrinsically disordered proteins play diverse roles in cell signaling.” Cell communication and signaling : CCS vol. 20,1 20. 17 Feb. 2022.
10. 10.- Buljan, Marija et al. “Alternative splicing of intrinsically disordered regions and rewiring of protein interactions.” Current opinion in structural biology vol. 23,3 (2013).
11. 11.- Metallo SJ. Intrinsically disordered proteins are potential drug targets. Curr Opin Chem Biol. 2010 Aug;14(4):481-8. doi: 10.1016/j.cbpa.2010.06.169. Epub 2010 Jul 2.
12. 12.- Saurabh S, Nadendla K, Purohit SS, Sivakumar PM, Cetinel S. Fuzzy Drug Targets: Disordered Proteins in the Drug-Discovery Realm. ACS Omega. 2023 Mar 8.
13. 13.- Chen J, Kriwacki RW. Intrinsically Disordered Proteins: Structure, Function and Therapeutics. J Mol Biol. 2018 Aug 3.