A calorimeter at atomic resolution

Informations projet

CLAR

N° de convention de subvention: 101087826

DOI

10.3030/101087826

Date de signature de la CE 24 Février 2023

Date de début 1 Juin 2023

Date de fin 31 Mai 2028

Financé au titre de

• European Research Council (ERC)

Coût total

€ 2 955 000,00

Contribution de l’UE

€ 2 955 000,00

Coordonné par

UNIVERSITAT WIEN
Austria

Periodic Reporting for period 1 - CLAR (A calorimeter at atomic resolution)

Période du rapport: 2023-06-01 au 2025-11-30

Résumé du contexte et des objectifs généraux du projet

Proteins are essential molecules that carry out nearly all functions in living organisms. They work by interacting with other molecules, such as small compounds, metabolites, or drugs. Understanding these interactions at the atomic level is crucial for biology and medicine. Today, most available techniques provide mainly static pictures of proteins, making it difficult to understand why some interactions are strong, others are weak, and why certain drugs bind only to specific sites. These limitations restrict progress in fields such as drug design, enzyme engineering and the study of diseases driven by malfunctioning proteins.

The CLAR project aims to overcome this gap by developing a new way to measure the thermodynamic forces that drive molecular interactions, and to do so with atomic resolution. The project uses advanced nuclear magnetic resonance (NMR) spectroscopy combined with computational modelling to create an “atomic calorimeter”, a tool capable of revealing how energy and entropy contribute to protein structure, stability and binding. By doing so, CLAR seeks to provide insights that cannot be obtained from structural data alone.

The overall objectives are:
• to build a framework that links NMR observables to thermodynamic quantities at the level of individual atoms;
• to map free-energy landscapes and identify regions of proteins that control folding, flexibility and binding;
• to apply this approach to proteins and protein–ligand complexes to reveal how dynamic features influence biological functions;
• to open the way for more rational and quantitative strategies in drug discovery.

The expected impact of the project is significant. A deeper understanding of atomic-level thermodynamics will help scientists interpret biomolecular behaviour more accurately and could support the development of new drugs, especially for targets that are difficult to study with existing methods.

Travail effectué depuis le début du projet jusqu’à la fin de la période considérée dans le rapport et principaux résultats atteints jusqu’à présent

During the project, the CLAR team developed a comprehensive experimental and computational workflow that allows free-energy mapping of proteins using multi-temperature and multi-field NMR spectroscopy. This includes an automated analysis pipeline that extracts atomic-level thermodynamic information from NMR chemical shifts and other observables.

The project successfully produced the first two-state ensemble structures derived directly from free-energy landscapes. These structures reveal how proteins populate different conformations and how these conformations contribute to stability and function. Through these analyses, the project identified entropic hotspots—regions that play a key role in flexibility—and dynamic reservoirs in disease-relevant proteins such as KRAS. These findings highlight features that cannot be recognised through traditional structural approaches.

The methodology was then extended to protein–ligand complexes, demonstrating that atomic entropic contributions to binding can be measured experimentally. This represents a major advance for understanding molecular recognition and for guiding the design of compounds that exploit hidden thermodynamic features.

From a methodological perspective, the project also developed new metabolic labelling strategies, enabling precise detection of specific residues that are critical for probing folding and interaction energetics. A unified theoretical–experimental framework was established to link NMR signals with enthalpic and entropic parameters at atomic resolution.

The work to date confirms the feasibility and power of the CLAR concept. Several scientific manuscripts reporting these results are in advanced preparation, and the new techniques are already being adopted for studies of protein dynamics and ligand binding.

Progrès au-delà de l’état des connaissances et impact potentiel prévu (y compris l’impact socio-économique et les conséquences sociétales plus larges du projet jusqu’à présent)

The CLAR project moves the field of structural biology from a static picture of molecules towards a thermodynamic and dynamic description at atomic resolution. This shift represents a major advance beyond current state-of-the-art techniques, which typically provide structures but not the energy landscapes that govern biological behaviour.

Key results include:
• the first experimental approach to derive atomic enthalpy and entropy contributions across entire proteins;
• a new way to identify dynamic and entropic features that influence folding, allostery and molecular recognition;
• the extension of this analysis to protein–ligand systems, opening new possibilities for drug design based on thermodynamic fingerprints;
• methodological innovations that integrate NMR spectroscopy with computational modelling in a unified, largely automated pipeline.

These developments have strong potential impact. Understanding thermodynamics at this level can guide the design of more effective and selective drugs, particularly for targets that are highly dynamic or difficult to crystallise. It can also support the engineering of proteins with improved stability or altered binding properties.

For further uptake, key needs include the expansion of computational tools, dissemination of the modelling framework, validation across diverse biological systems and potential integration with pharmaceutical screening pipelines. The project already provides the theoretical basis, experimental evidence and methodological tools needed for such next steps.

Overall, CLAR delivers a transformative approach that goes beyond structural characterisation to reveal the energetic principles that shape biomolecular function. This lays the foundation for broad applications in biology, chemistry and drug development.

Atomic resolution calorimeter