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Tuo Wang (left) and Fred Mentink-Vigier study the cell walls of fungi. Photo: Stephen Bilenky
harness this technology for an energy source, it would be fantastic. And bacteria have been doing this for millennia.” When Griffin tried conventional NMR to study how ions were crossing cell membranes during this photocycle, he got a lot of noise in his data. It’s like trying to listen to a song on your car radio when the station is out of range: You’re picking up signals, but the static makes it hard to hear the music. “We did a lot of experiments without DNP,” remembered Griffin. “But it became very apparent very, very quickly that if we had a big signal-tonoise enhancement technique, and we could do spectroscopy at low temperatures, we could learn a lot more about the system.” That turned out to be a pretty ambitious “if.” MAS DNP didn’t exist yet. He would have to build it. Some doubted it could be done, but Griffin was in it for the long haul. “He was the lone voice saying it was going to work out and it was going to be great for everyone,” recalled Joanna Long, a former graduate student of Griffin’s. There were plenty of hurdles, including building a gyrotron to generate the frequency of microwaves his set-up required. But eventually it paid off with an instrument that could detect the subtle signals of the carbon, nitrogen and hydrogen he was trying to map. Today, Griffin’s lab at the Francis Bitter Magnet Laboratory boasts several home-built DNP systems. Now when Griffin examines the bacteriorhodopsin 10
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photocycle using MAS DNP, very cold temperatures and lasers of different colors, he can capture incredible details at specific stages that shed light on how hydrogen crosses cell membranes. “What we’re interested in is how this proton pump works, how this inward-directed hydroxyl pump works,” Griffin said. “And in order to understand that, we have to trap various intermediates in this photocycle.” The MAS DNP improved the signal-to-noise ratio by a factor of 75, Griffin said, making the experiment thousands of times faster. “It’s an enormous time savings,” said Griffin, who also uses MAS DNP to study the amyloid protein fibers associated with neurodegenerative disease. “You just couldn’t do these experiments without this increase in sensitivity.”
Prepping samples isn’t simple While dozens of commercial instruments are in use worldwide for dissolution and MAS DNP, a third DNP technique, Overhauser DNP, has proven a tougher puzzle. But when those pieces come together, it will fill a vast void by allowing scientists to study molecules in liquids at low concentrations. Several teams around the world are developing the technique. At the National MagLab, physicist Thierry Dubroca has devoted several years to assembling a machine and getting it to work. But that may have been the easy part.