ATP is the stuff of life. Without it, cell communication shuts down, muscles freeze, and anything requiring ready energy (which is to say, darn near everything) stops. For biologists, it’s a critically important molecule to understand, but most of its secrets remained locked up until one person, who was very nearly shut out from scientific research altogether, thought of elegant new ways of applying the technique of nuclear magnetic resonance to probe its depths.
Mildred Cohn (1913 – 2009) was the daughter of a rabbi and grew up in an environment of culture and education, her teen years spent in a Yiddish collective where learning and social justice were the orders of the day. The supportive environment, combined with her raw talent, allowed her to graduate high school at 14 and college at 18. She managed to finance a year of graduate studies at Columbia by working at a department store and finished her Master’s degree in physical chemistry in that short time, but then she was brought to a hard stop by the institution’s refusal to give assistant teaching positions to women. Without that position, she couldn’t afford to continue her PhD studies, and so she had to take a job in industry.
She found work at NACA as a fuel injection chemist, the only woman in a group of seventy. While there, an administrator was horrified to see her working in the lab alongside the men. Concerned that the presence of a woman would fatally distract his male researchers, he ordered her out of the lab. She was allowed to design experiments, but then she had to hand them off to a male colleague to actually perform.
Yeah, I know.
With no chance at promotion, and cut off from the lab work at which she excelled, she left after two years, returning to Columbia and the lab of Nobel laureate Harold Urey, who told her from the outset that she would be on her own, as he spent precisely zero time advising his graduate students.
She made a name for herself early on as not only an ace chemist, but as a master of machinery. Without a budget, she had to build whatever equipment she needed, and constructed a mass spectrometer from scratch at the labs of both Urey and her next advisor, Vincent du Vigneaud, who was reluctant to hire a woman but relented when he couldn’t find anyone else with her mechanical-chemical skill set.
The work that defined her career, however, didn’t come until 1938 when she and her physicist husband moved to Washington University in St. Louis. There, she had the opportunity of working in Gerty Cori’s lab, where field-defining research on enzymes and the major biochemical cycles was being carried out. Cohn decided to work on ATP, the molecule that stores up the energy our cells need to go about their business. She investigated its structure, how it is broken apart in different metabolic reactions, and started unraveling the intermediate steps in the great energy-generating processes of oxidative phosphorylation that power all living things.
Her tool of choice was nuclear magnetic resonance, or NMR, a technique that offered major advantages over the mass spectrometry of her earlier research. Mass spectrometry was a useful but destructive process that used electron bombardment to break apart molecules. By measuring the charge to mass ratio of the resulting pieces, one could then determine the compound’s molecular formula.
NMR, by contrast, was far less invasive and as such held out the promise of much more detailed molecular information. It’s a neat process, a sneaky hack on nature, that deserves a closer look. If a nucleus has an odd number of protons or neutrons, it has one of two possible spin states, which we call +1/2 and -1/2. +1/2 nuclei align with an applied magnetic field and -1/2 align opposite one. Because of this, +1/2 and -1/2 nuclei have a tiny difference in energy when you put them in a magnetic field, a difference that can be probed by a radio wave with a frequency corresponding to that energy difference (just rewrite the old high school formula E = hf as f = E/h and you’ll get the radio wave that should do the job!)
So, instead of blasting molecules apart, we can just gently nudge them with relatively low energy electromagnetic waves. More importantly, the frequencies required to probe a nucleus change in response to the other atoms around that nucleus. Some atoms are way electron hungry and will draw the electrons of neighboring atoms towards them. A hydrogen that’s connected to a hungry atom like oxygen or fluorine will have less electron cover as a result, and that will change the recorded frequency of the Hydrogen, giving us information not only about the presence of a hydrogen atom, but also about what it’s connected to! Snazzy!
As a matter of fact, there is a ton of structural information lying in that frequency. By watching the frequency shift at differing molecular concentrations, you can find out how much hydrogen bonding there is, and therefore know something about the intermolecular forces that govern so much of a substance’s physical properties. Pi bonds (the bonds responsible for double and triple bonding) have tell-tale frequency impacts as well. The list goes on and on, and Mildred Cohn was a master of this subtle new method.
One of my favorite pieces of her work was how she used O-18 to find out how the oxygens in ATP move around in different reactions. O-18 has an even number of protons and neutrons, and so can’t be measured directly by NMR, but each O-18 has a marked impact on the phosphorous atom it’s connected to, and that’s an atom we can use NMR on. So, by keeping an eye on how the phosphorous frequencies hopped about, she was able to tell what was happening in the hidden world of the oxygens, and thereby detail the complicated restructurings that ATP often goes through.
ATP is made up of adenine and ribose (don’t worry about it) connected to three phosphate groups, PO4 3-. Each of those has a negative charge, meaning they reaaaaally don’t like being smooshed together, and that you can harvest some energy whenever you pluck one off. That mechanism basically runs our lives, and Cohn used NMR to determine where the cleavages happen, how enzymes based on different metallic ions interact with it, how the molecule reorients itself during different reactions, and pulled back the curtain on the hidden intermediate processes that our cells use to make ready energy from the products of photosynthesis.
It was outstanding work and, astoundingly, she did not receive a full professorship for it until 1960, twenty two years after her work with the Coris began. Better times, and the explosion of MRI technology, brought further recognition, and she was awarded the National Medal of Science in 1983 and the Chandler Medal in 1986. She was ambitious, demanding, brilliant – the author of 160 papers, and a pioneer of a field that changed how we diagnose illness. She died at the age of 96 in 2009, but her masterful work continues to save lives and prod the fuzzy edges of biochemistry, where structure is king and the unknown runs deep.
FURTHER READING: NMR and Biochemistry: A Symposium Honoring Mildred Cohn (1979) features a brief introduction from Carl Cori talking about her work in the Cori lab, and is followed up by a summary by Cohn herself of some of her accomplishments in NMR. The rest of the book is made up of a series of papers by other NMR scientists who don’t deal directly with Cohn, so your choice if you want to buy a 400 page science volume for 26 pages of Cohn content. I did. On the internet, there’s a neat 18 minute video put out by the Chemical Heritage Foundation that you can find here which focuses primarily on her life and struggles for professional recognition.