Studying proteins @ work in cells using magnetic resonance In all cells, proteins are essential building blocks that are needed to execute and change cellular functioning. For a long time, it has been known that many proteins can change their conformation but if and how this occurs inside the cell has been difficult to follow. This project will develop a Magnetic resonance (NMR) -based approach to directly probe at atomic scale how dynamic proteins change conformations during cellular function.

Extremely drug-resistant pathogenic bacteria, so-called superbugs, are a major cause of human death. Superbugs are often protected against antibiotics by a massive cell envelope, a wall-like structure that surrounds them like a thick armour. On the other hand, since the envelope is essential, antibiotics that directly target the bacterial armour could make powerful drugs. Unfortunately, the mechanisms how these drugs work are not well understood due to technical challenges. This project will develop concepts and technologies to study these ‘envelope-targeting antibiotics’ in detail, focusing on antibiotics against the most dangerous superbugs: Gram-negative bacteria and Mycobacteria.

Structural biology is vital in understanding details of biological molecules, shaping our knowledge of how cells work with applications in medicine and more. During COVID-19, the WeNMR computational services have been crucial in modelling viral interactions. To continue this work, continued access to the Dutch high throughput computing grid resources is needed, supporting global research and collaborations in structural biology.

Pioneering chromatin: interaction of pioneer proteins with native, genomic nucleosomes Nucleosomes are the building blocks of chromatin and essential to swich off genes that are not needed by a cell. This way the identity of a cell, e.g. a liver vs. neuron cell, is determined. So-called pioneer proteins can however bind nucleosomes to activate these genes, thereby transforming cell types and even changing “adult” cells to stem cells. In this project, we will use NMR spectroscopy and biochemical techniques to unravel how pioneer proteins bind native, genomic nucleosomes. This will be useful to improve cell fate conversions and regenerative therapies.

Membrane proteins code one third of all genes in all organisms. They are involved in any known metabolic or signalling pathway and constitute half of all drug targets. To date, since membrane proteins are inappropriate for conventional structure determination techniques, only one percent of all known protein structures account for membrane proteins. Moreover, membrane proteins act in dynamic interplay with their membrane environment, which is hitherto poorly understood on a molecular base. This crucial protein?-membrane dialogue shapes protein structure, determines protein orientation relative to the membrane (known as topology), modulates protein?-protein communication by steering protein localisation and assists protein folding. Together, structure, topology and protein-?membrane interplay comprise the supramolecular structure, i.e., the complete picture of a membrane protein. This proposal aims to study the supramolecular structures and functions of large integral membrane proteins using a strongly complementary joint-approach of solid state Nuclear Magnetic Resonance (ssNMR) and Molecular Dynamics (MD) simulations. SsNMR allows investigating membrane proteins in their native environment at atomic-resolution. Large membrane proteins, however, only give access to spectra of low quality. This shall be solved by tailoring ssNMR methods to work at optimal sensitivity and resolution provided by ultra-high magnetic fields and spinning frequencies, proton-detection and dynamic nuclear polarisation. SsNMR information will be complemented by MD simulations to refine and validate protein structure and to establish protein topology by back-calculating ssNMR observables (like the protein water-access) over MD trajectories. I will particularly focus on protein-membrane interactions by virtue of combined ssNMR-MD investigations with ssNMR experiments especially designed to be correlated to MD trajectories. This approach shall be applied to model membrane proteins with known crystal structures (ion channel KcsA, photosensor ASR) and employed to study beta-barrel assembly machinery BamA (88 kDa), which is a key player in membrane protein folding of unknown high-resolution structure.