Dynamic Nuclear Polarization (DNP)

Biomedical Research Utilizing Dynamic Nuclear Polarization

Principal Investigator: Joanna Long
Program Director: Frederic Mentink-Vigier

The development of dynamic nuclear polarization (DNP) to increase the sensitivity of NMR by 1-2 orders of magnitude is profoundly impacting the field of NMR. We address making DNP an enabling technology for biomolecular solid-state NMR by maximizing the promise of DNP through hardware development, synthesis of needed chemical reagents and optimizing sample preparation strategies.

Advances in all these areas will gain the needed sensitivity and resolution for leading edge biomedical research applications and transition DNP MAS solid-state NMR to a technique that is more widely accessible to the structural biology community as a whole. Through partnering with Driving Biomedical Research projects, we are developing and deploying technology in this TR&D to address biomedical research problems in the areas of membrane protein structure determination, amyloid biogenesis in bacterial biofilms and in neurodegenerative diseases, protein assembly in viruses, and enzyme catalysis.

• Aim 1: Develop fast MAS and extend VT range for MAS-DNP ssNMR probes. This aim focuses on hardware improvements to support biomolecular MAS-DNP ssNMR applications. This includes fast MAS to further improve resolution and sensitivity, lowering temperature to improve polarization enhancement, developing a suite of probes enabling users to interrogate different NMR isotopes as dictated by their needs, and manipulating the μw beam using quasi-optical approaches enabled at high fields.
• Aim 2: Develop sample protocols maximizing DNP enhancement of biomolecular samples. The effectiveness of MAS-DNP for polarizing molecular species is dependent on many parameters that have over the years been slowly elucidated. The electron spin properties, strength of the coupling between the radicals, a biradical’s geometry, the nature of the chemical species, and the distribution of PAs within a sample all determine the efficiency of the nuclear hyperpolarization. Sample integrity and PA distribution are dependent on their nature but also on how a sample is prepared and frozen. This aim focuses on recently developed biradicals and optimizing sample preparation strategies for different types of biological samples.
• Aim 3: Optimize MAS-DNP ssNMR approaches for characterizing biomolecular samples. The sensitivity of MAS-DNP NMR opens new horizons for utilizing low gamma / quadrupolar nuclear spins to understand biological chemistry in addition to enabling structural characterization of very challenging samples. While early MAS-DNP studies suggested a degradation in ¹³C/¹⁵N resolution, relative to conventional ssNMR, we demonstrated through our TR&D that we can achieve high resolution even under DNP conditions. With the improved capabilities of the MAS-DNP NMR probe we developed, we are now poised to further improve resolution and greatly expand the range of NMR beyond commercial DNP probes with a ¹H/¹³C/ ¹⁵N configuration.

Associated Driving Biomedical Projects (DBPs):

• RNA Recognition by Intrinsically Unstructured Proteins; Robert Silvers, Florida State University
• Structural and proton dynamics of pyridoxal-5′-phosphate dependent enzymes; Leonard Mueller, UC Riverside
• Molecular organization and drug responses of fungal cell walls, Tuo Wang, Louisiana State University
• Functional amyloid formation in Streptococcus mutans; L. Jeannine Brady, University of Florida
• In situ mapping energy landscapes of biological macromolecules and their complexes, Matt Eddy, University of Florida

Associated Collaboration & Service Projects (CSPs):

• Structure and Dynamics of Self-Assembled biomolecules; Bo Chen, University of Central Florida
• Mechanistic Investigations of Host Defense Metallopeptide Interactions with Bacterial Cell Membranes; Myriam Cotten, William & Mary
• Functionalized gold nanoparticles; Terry Gullion, West Virginia University
• ⁴³Ca, ²⁵Mg NMR biomaterials; identification of interfacial bonding environments in functional nanomaterials and biomaterials using high resolution solid state NMR at (ultra)-high fields; Christian Bonhomme, Condensed Matter Chemistry Laboratory, Sorbonne University, Paris, France
• Protein molecular structure, conformational dynamics, and inter-protein interactions in human health and disease, Dylan Murray, University of California, Davis
• Dynamic Nuclear Polarization of Membrane Proteins in Lipid Bilayers by Solid-State NMR, Alexander Nevzorov, North Carolina State University
• ssNMR Structural Analysis of Oligomeric Alzheimer’s Beta-Amyloid Peptide; Anant Paravastu, Georgia Tech
• Structural Studies of FMN domain interactions with cytochrome-P450; Ayyalusamy Ramamoorthy, University of Michigan
• Interaction bones/cartilage with extra-cellular medium; Neeraj Sinha, University of Miami Health
• Structure and signaling-related changes at protein interfaces in chemotaxis receptor arrays; Lynmarie Thompson, University of Massachusetts-Amherst
• Dynamic structures of lipid-membrane protein complexes via ssNMR; Benjamin Wylie, Texas Tech University

Technology Partnership Projects (TPPs):

• Rapid freeze quenching, Affinity biradicals, Relaxation sequences; Ann McDermott, Columbia University
• Preparation of viral capsid assemblies, Fast MAS and ¹⁹F sequences; Tatyana Polenova, University of Delaware
• Mechanochemical methods for ¹⁷O-enrichment of amino acids; Danielle Laurencin, Institut Charles Gerhardt, University of Montpellier, France
• DNP sample preparation and cell/membrane viability; Kendra Frederick, UT Southwestern

To inquire about starting a collaboration including establishing a DBP, CSP, or TPP, please contact principal investigators for this TDP: https://nmrprobe.org/people/leadership/.