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Content archived on 2024-04-30

NMR structures of membrane proteins, complexes and lipid assemblies; a dedicated wide-bore ultra-high-field MAS NMR spectrometer for biological research


The objectives are to establish and to validate ultra high field Magic Angle Spinning NMR technology required for comprehensive structure determination using multidimensional correlation spectroscopy. This involves the construction of a fully operational prototype spectrometer with dedicated high-speed MAS probes, operating at ultra high field ( 750 MHz) with excellent long-term stability, and the construction of the associated ultra high field wide-bore magnet, in a unique partnership between scientific key laboratories and the only major NMR manufacturer in Europe. A demonstration phase will overcome the techno-economic uncertainties associated with the implementation of the spectrometer and the new technology. The novel instrumentation will be available to all biological solid state NMR users in Europe. The user exploitation will prove the concepts for MAS NMR structure determination, and will demonstrate routine specification of ligand-protein interactions for biomolecules with the desired functions (e.g. drugs, hormones and membrane protein receptors) using multi-spin and specific isotope labeling of genuine subcellular entities, as defined in Area 6.1 Structural biology, research tasks 6.1.1 6.1.2 (prominent) and 6.1.3. The long-term goal is to establish flexible technological means to obtain comprehensive molecular information, at atomic resolution, for widely different types of molecules or subjects, and to relate this to function, eg. for an essentially unperturbed membrane protein receptor when in the membrane. A preparation has confirmed the feasibility of the novel instrumentation. Multidimensional homo-nuclear and hetero-nuclear test-experiments using multiple 13C labeled and 2H diluted samples have been performed to determine the instrument requirements with respect to long term instrument stability, temperature range, rf power requirements, magnet specifications, optimization of existing rf technology for ultra high field MAS, and a structure was partially resolved. This proposal provides a detailed plan on how to combine the most advanced knowledge from the industrial and scientific key-players in Europe into a multi-disciplinary enterprise to design unique and widely applicable MAS NMR technology for biological studies, including a dissemination strategy of how to make the new instrumentation accessible to all biological solid-state NMR users in Europe, in the form of a facility that will be open for all, and taking into account crucial requirements in the field of isotope chemistry and molecular biology methods for membrane protein labeling, reconstitution and NMR development. Economic benefits will accrue for health related industries from gaining urgent knowledge of ligand (eg: drug)-receptor binding site. With access to such information rationale design of biomolecules with the desired functions (eg. drugs) becomes a reality. Biotechnological benefits will accrue from forging the pathway for the determination of integral membrane protein structure. NMR manufacturing industry will benefit from a new product brought into production for a potentially developing market.

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Rijksuniversiteit Leiden
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2333 RC Leiden

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Participants (12)