Studies in mice and human stem cells demonstrate that the genome-editing technique CRISPR can correct disease-causing mutations.
JoAnne Stubbe's determination has unlocked the secrets of ribonucleotide reductase.
February 1, 2007|
After receiving her PhD in chemistry from the University of California at Berkeley in 1971, JoAnne Stubbe did a very brief stint as a postdoc at UCLA, where she worked on synthesizing LSD from tryptophan. She then decided to step up to the chalkboard. "Both my parents are teachers, so I thought that's what I wanted to do," she says. She accepted a teaching position at Williams College, a small liberal arts institution. But she soon realized that "teaching is not what it's cracked up to be. You always have a few kids who love science." Most of what she heard, however, was complaining. "The longer I was there, the more I realized I wanted to do research."
That realization sent Stubbe to Brandeis University, where she did a second postdoc with Bob Abeles. There she learned the art and science of producing mechanism-based enzyme inhibitors, a skill she would exploit to great success later in her career. Although the experience convinced Stubbe that she wanted to devote herself to research, her love affair with the bench actually began when she was a student in high school.
"From day one I really loved the laboratory," says Stubbe, now a senior scientist at Massachusetts of Technology, where she's been since 1987. "The color changes, the crystals, the smells. I just liked mucking around in the lab. Seeing things change, making observations. And I still like that. There's nothing that replaces the thrill of making a discovery yourself. In fact, many of the discoveries that got me where I am, I made with my own hands."
"I remember standing there watching counts coming off the counter with her," says Georgianna Harris of Merck, one of Stubbe's first graduate students. "Day or night, it didn't matter what time it was. We were so excited about what was going on."
At that time, what was "going on" were experiments designed to explore the intricate molecular maneuvers that underlie the activity of ribonucleotide reductase, an enzyme that Brian Hoffman of Northwestern University says "makes DNA-based life possible." Ribonucleotide reductase converts ribonucleotides into deoxyribonucleotides, the building blocks of DNA. "Without it you don't have any higher-order life," says Hoffman. Or, as Stubbe puts it, "without it, you're dead."
Those round-the-clock efforts (for which Stubbe reportedly kept a bed in her office) allowed her to make great strides in unraveling the mechanism by which ribonucleotide reductase generates deoxyribonucleotides, a problem she continues to work on in her lab at MIT. "Before her first contribution to that work, there was no chemical mechanism that could be written," says Perry Frey of the University of Wisconsin. "After her first experiments, there was. She revolutionized that field with her first two papers."
Those early, groundbreaking experiments were carried out in the late 1970s and early 1980s while Stubbe was an assistant professor, first at Yale (from 1978 to 1980), then at the University of Wisconsin, where she remained until 1987. (Including her years at Williams, Stubbe was an assistant professor for 12 years, "which would give most young people a nervous breakdown these days," she laughs.) The chemistry Stubbe was trying to understand involves replacing the hydroxyl group at the 2' position of the ribonucleotide's sugar with the hydrogen found in deoxyribonucleotides. "All you're doing is cleaving a carbon-hydroxyl bond and forming a carbon-hydrogen bond," she says. "It looks like it's pretty simple, but the chemistry is amazingly complex."
"She knew as an organic chemist that that reaction is impossible," recalls Frey. "There's no established chemistry that would allow it to happen. So she got to thinking about how nature might get around that problem. And she realized that if the carbon-hydrogen bond at the 3' carbon were broken, an intermediate would be formed that might allow the chemistry at the 2' carbon to occur." In other words, the first step in reducing the ribonucleotide involves cleaving a bond at the carbon that's next to the carbon where the "chemistry" ultimately occurs. Such a reaction "was just totally unprecedented in biochemistry," says Judith Klinman of the University of Berkeley.
"It was like a light coming on in a dark room," adds Frey. "JoAnne has done many things since then that are comparably important. But that was revolutionary. As soon as she did that she became a leader in the field."
The work, which Stubbe says earned her tenure, was not easy. To do the proper experiments, Stubbe used nucleotides that carried a heavy isotope at specific positions, so she could follow what was happening to each part of the molecule. First she had to synthesize those compounds, a process that Frey describes as "demanding and not something to be undertaken lightly."
"They were difficult experiments," agrees Wallace "Mo" Cleland, another former colleague from the University of Wisconsin. "But JoAnne is a very good chemist. Nowadays a lot of biochemists are molecular biologists and all they do is use kits they buy from Sigma. But JoAnne is an excellent organic chemist and an imaginative scientist. She can make what she needs and she's always on the cutting edge of things."
Stubbe's work has led to the development of mechanism-based inhibitors, such as gemcitabine, that are being used as drugs to fight pancreatic cancer and non-small-cell lung carcinoma. Battling tumors was not Stubbe's original goal. "I've never been one who's driven by something practical," she says. "I focus on trying to understand basic biology, which I find really interesting. It just so happens that almost all of the systems we work on have something practical out the other side."
"JoAnne doesn't want to change the world or find cheaper ways to do something. She's just curious about how nature gets things done," says Barry Sharpless of Scripps. "She can identify the unforeseen gem of reactivity that, no matter how crazy it looks, turns out to be the one nature is using to accomplish the impossible. And when she's through, everyone can see: 'Oh yeah, it's not magic, it's chemistry.'"
Stubbe is determined when it comes to her science. Witness the fact that she is still working on the mechanism of ribonucleotide reductase after some 30 years. "That's an extraordinary thing," says Peter Lansbury of Harvard Medical School. "A lot of people are very satisfied making contributions in various areas and then getting out. JoAnne doesn't want to leave it at 'a great contribution' and then let other people push it over the goal line. She wants to get it 100%. She's totally focused on solving a problem and is not willing to stand down."
In the case of ribonucleotide reductase, such determination has led Stubbe to constantly learn new techniques and new methods for chasing down the free radicals that allow the enzyme to do its job. Because of their extreme reactivity, these fiery chemicals tend not to stick around long enough to be captured with experimental ease. To follow the radicals, Stubbe has adopted "all manner of exotic spectroscopic methods," says John Kozarich of ActivX Biosciences in San Diego. "She's fearless when she wants to answer a question," adds Klinman. "After she defines a problem, she's relentless in getting the experimental data to find out if her hypotheses are right. She defines the most appropriate experiments, finds the right collaborators, and moves ahead to get the experimental evidence."
"She'll go wherever she needs to go to find the right technologies to answer her questions," says Kozarich, who was Stubbe's colleague at Yale. John Gerlt of the University of Illinois, another former Yalemate, agrees. "JoAnne is not afraid to go where the problem takes her. As you move along with a problem, you look at it in different ways and apply new techniques. She's also not afraid to collaborate," says Gerlt. Working with Stubbe "is a gas," says Hoffman. "She's so inquisitive, so intellectually fearless, that to play on her court you'd better be at the top of your game. And that's fun and exciting."
"There are few people as dedicated as she is to fundamental science," says Tony Sinskey, a colleague and collaborator at MIT. "For me, she's one of the best colleagues I've ever had at MIT."
For her part, Stubbe enjoys exploring new methodologies and establishing new scientific relationships. "It's a continual process of learning, which is why I like my job so much." Stubbe will visit a lab, pick up a new technique, and then bring it home or set up a collaboration. "Over the years I've had a lot of outstanding collaborators, which is part of what makes science fun," she says.
It also helps her get the answers she craves. For example, one piece of the ribonucleotide reductase puzzle regards the movement of the electron that reduces the nucleotide from one side of the protein to the other. "The actual chemistry is done in one subunit, where you cook the substrate to make the product," says Hoffman. "And the fire that does the cooking has to be lit. But it turns out that the pilot light that lights the fire is a long way away," 35 Ångstroms away, to be precise. That's much too far for an electron to leap in a single bound. Using several techniques for incorporating modified, unnatural amino acids into a protein - one of which she picked up in Peter Schultz's lab at Scripps - Stubbe identified a tyrosine residue that serves as a pit stop for the traveling electron. That was three years ago, and Stubbe is still searching for other electron-friendly residues that might line that path, in her continuing quest to understand ribonucleotide reductase completely.
"Her whole life is science," says Sinskey. "She'll drop anything to sit down and have a discussion about your science or her science," says Jamie Williamson of Scripps, who was formerly a junior faculty member in Stubbe's department at MIT. "That's what was most refreshing about her. Everything for her is about science. She's just raw science."
Stubbe is not the only one in her group who likes to sink her teeth into a juicy new piece of data. Her dog McEnzyme regularly attends lab meetings, where he loves to watch the laser pointer move across the slides. "Except for JoAnne, he's often the most attentive one in the room," says MIT colleague Cathy Drennan. "It's fun to watch both JoAnne and her dog enjoying the presentation and getting excited about science."
Although Stubbe admits that science is her passion, she says she's currently thinking about the idea of retirement. Perhaps she's hoping to have some time to read through the stacks of journals that are heaped around her office. "I like to read journals to see what the next great scientific breakthroughs are," she says. "Of course, nobody can believe that I won't die with my boots on."
"Science is her life. It's what she does. I can't imagine her willingly retiring," says Williamson. Drennan agrees. "I don't think people will let her escape. I don't know what would happen to us if she left."