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Electron microscopy of enzymatic pathways in polymer-stabilized nanodiscs
M.Sc. Janson Kevin
EU - ESF Sachsen-Anhalt ;
EUROPÄISCHE UNION - ESF -  Europäischer Sozialfonds
Electron microscopy of enzymatic pathways in polymer-stabilized nanodiscs
Part of ESF "AGRIPOLY" graduate school
Background: Electron microscopy is a powerful tool for studying enzymatic processes in various biological contexts. In particular, understanding the structural organisation and interactions of proteins within lipid bilayers and the role of lipids is crucial for deciphering complex biological processes. This research project focuses on the use of polymer-stabilised nanodiscs as a versatile model system to study such pathways, with particular emphasis on the myelin sheath in the context of neurodegenerative diseases.
Nanodiscs are polymers with the ability to interact with membranes and form flat, disc-shaped biomolecules with defined diameters. Their application in membrane protein research has revolutionised the field of membrane protein complex purification and structure determination, especially in combination with single particle cryo-EM technologies. In this project, we aim to develop nanodiscs of different polymers and solubilise liposomes made of artificial lipid mixtures that may or may not encapsulate membrane proteins. These lipid mixtures can simulate the properties and physicochemical composition of cell membranes, for example those of mitochondria or plasma membranes. By using different polymers and lipids, we will gain insights into the determinants of nanodisc formation. In addition, we will investigate nanodiscs that form after solubilisation of native membranes and characterise them using a variety of biophysical methods, including cryo-EM. Ultimately, our goal is to develop tools to structurally characterise native membrane complexes at high resolution. This result will provide a novel insight into the structure of membrane complexes with minimal biochemical intervention and thus come closer to understanding their natural-like function.
Preliminary work: Previous publications have laid the foundation for this project by presenting myelin-mimicking nanodiscs made of different amphiphilic copolymers, such as styrene/maleic acid (SMA) and styrene/maleimide-sulfobetaine (SMA-SB). These nanodiscs have been shown to effectively dissolve myelin-like liposomes and interact with myelin basic protein (MBP). Thus, they offer a promising platform to study protein-lipid interactions in a controlled environment. In addition, lipid analysis methods using thin-layer chromatography and high-resolution mass spectrometry were developed to improve the characterisation of lipid species in the nanodiscs.
Objectives: The main objectives of this research project are to further improve the understanding of enzymatic pathways in polymer-stabilised nanodiscs and to apply this knowledge to the study of the myelin sheath. Specific objectives include elucidating the structural and functional properties of lipid bilayers within nanodiscs, characterising the interactions between proteins and lipids, and exploring the potential applications of this platform to study neurodegenerative diseases.
Methods: The research uses a combination of techniques including electron microscopy, mass spectrometry and chromatography to analyse the structure, composition and behaviour of lipid bilayers in nanodiscs. The solubilisation efficiency of different copolymers in solubilising charged membranes will be investigated, and dynamic light scattering and negative-stain electron microscopy will be used to characterise nanodiscs. In addition, cryo-electron microscopy is used to study endogenous membrane-protein complexes in polymer-based nanodiscs with lipid layer.
Impact: Research on electron microscopy of enzymatic pathways in polymer-stabilised nanodiscs has significant implications, including advancing scientific knowledge in the study of lipid bilayers and protein-lipid interactions. It holds promise for understanding and potentially treating neurodegenerative diseases, provides innovative methods for lipid analysis and supports interdisciplinary collaboration. In addition, the research promotes sustainable and efficient research practices while contributing to global health and well-being by addressing critical issues in the field of neurodegenerative diseases.
Alignment with the Sustainable Development Goals : This research project has far-reaching implications for several Sustainable Development Goals (SDGs). It contributes to SDG 3 (Good Health and Wellbeing) by improving our understanding of neurodegenerative diseases and potential therapeutic targets. It is in line with SDG 9 (Industry, Innovation and Infrastructure) by developing innovative methods for studying biological processes. It also supports SDG 12 (Responsible Consumption and Production) by improving our ability to analyse and use biological materials efficiently and sustainably. The research is in line with several other SDGs, such as SDG 4 (Quality education) by promoting scientific knowledge and sharing results with the general public, and SDG 17 (Partnerships for the Goals) through potential collaborations in the fields of medicine, chemistry and materials science. In addition, the development of sustainable and efficient methods contributes to SDG 13 (Climate action) by reducing resource use in research, and has the potential to advance SDG 16 (Peace, justice and strong institutions) by contributing to the understanding of complex disease mechanisms and supporting medical research.

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