Progress in FY2017 was made in the following areas: (1) RAPID MIXING AND FREEZING TECHNOLOGY FOR SSNMR. We have designed and tested a new rapid mixing device, constructed from commercially available fluidic components and fittings. This device mixes two solutions in 10 milliseconds or less, and is compatible with flow rates, volumes, and viscosities that are relevant to our low-temperature dynamic nuclear polarization (DNP) experiments. Initial experiments on the folding of tetrameric melittin, initiated by rapid dilution of a urea-containing solution with urea-free buffer, demonstrate the functionality of this device when combined with our rapid freezing apparatus developed in FY2016. Various experiments are planned, including studies of the initial stages of self-assembly of the HIV-1 capsid protein to form an ordered protein lattice, which will involve rapid mixing of an unassembled capsid protein solution with high-salt buffer to initiate the lattice assembly process. (2) MRI MICROSCOPY. We have obtained the first high-resolution 3D magnetic resonance images of test samples at low temperatures (15-30 K) range. The signal-to-noise enhancement that arises from enhanced nuclear spin polarizations at low temperatures allows us to achieve isotropic resolution of 2.8 microns over a total volume of 1.5 nanoliters. This represents an approximately 30% improvement over the best MRI resolution previously reported in the literature. A publication to describe this work is currently in preparation. (3) VERDAZYL RADICALS FOR DNP. We have demonstrated that large NMR signal enhancements can be achieved by cross-effect dynamic nuclear polarization in frozen solutions using soluble stable free radical dopants based on the verdazyl moiety. Results with verdazyl are comparable to those obtained with standard nitroxide-based dopants. Calculations suggest that verdazyl-nitroxide biradical compounds may be more effective than any previously reported DNP dopants. A manuscript describing this work has been submitted for publication.
Showing the most recent 10 out of 22 publications
