Eileen Jaffe has spent a long career as a professor in the Molecular Therapeutics program at Fox Chase Cancer Center, one of the leading cancer centers in the US and one of the oldest NCI designated Comprehensive Cancer Centers (it formally became part of the Temple University Health System about a decade ago). She received a PhD in biochemistry from the University of Pennsylvania in 1979, studying under Mildred Cohn as an NSF Graduate Fellow. She did her postdoctoral work with Jeremy Knowles in the Chemistry Dept. at Harvard (1979-1981), on an Individual NIH Post-doctoral fellowship.. She has done extensive work on protein structure, and was NIH-funded for many years.
Can you describe your background in science?
I’m a classically trained enzymologist—a biochemist. I was born in the 1950s, and had a penchant for science, and was pushed ahead in school (I was put into a program that did 4th, 5th, and 6th grade in two years). I was a Sputnik kid, and was never told “girls can’t do this.” After my BS, I briefly took a job working in industry as a chemist, and still got my PhD before I was 25.
It wasn’t until I was a post-doc at Harvard, and one of the faculty members saw me in the stock room, and handed me a paper to type because he thought I was a secretary—that I first thought, “Oh wait, maybe it is unusual for a woman to do this.” Yes, I was clueless!
I cannot know how much my experiences in science have to do with being a woman. There is no “control” for this question. I do know that when I put male technicians’ names on my papers, I had an easier time getting them published.
The implicit bias against women (in science) is very real. I was once on a search committee for a faculty member, and I was so impressed with the accomplishments on their CV, and I read the letters of recommendation and was shocked to find out it was a woman. In my mind, it was a man. Apparently, even I can impose implicit bias. I have to keep that in mind when I think about the pushback I’ve gotten at various points in my career.
In layman’s terms, what have you focused on in your scientific career?
Mostly, I have focused on questions of protein structure and function. Back in the 1970s, a guy named Anfinsen determined that if you take a protein, and do something to take out its structure, it can refold and regain its function. So it became a sort of dogma that each protein has one structure and one function. The huge field of bioinformatics is based on this fundamental principle that the sequence of a protein determines its fold and function.
When Prusiner first started looking into prions, and determined that proteins could be folded differently—well, that was Nobel-level work that couldn’t get funded at first. In the case of prions, there was one right fold and function (the Anfinsen principle), and a disease-ridden way of doing it (the amyloid-promoting alternate fold).
Around 2000, we stumbled on an observation that was almost unthinkable. For the protein we were studying, the assembly of identical protein chains came apart, the individual components changed shape, and the assembly came back together. It was not irreversible like prions, it just kept happening. And it was physiologically relevant, not a laboratory artifact. One assembly was “on” and the other assembly was “off”.
Now there are lots of reasons in biology where you might want to turn a protein on or off. We discovered there were small molecules that could regulate this on/off switch. These molecules were binding some place on the protein different from the active site. So often an alternate form of a protein that is not “on” is just assumed to be misfolded or an experimental artifact.
We had an octameric assembly (eight parts) that could come apart, change shape, and reassemble to a hexameric assembly (six parts). The shape change was not a refolding . . . just a repositioning of domains. And this repositioning could not happen in either the 6-part or the 8-part assembly. I remember in the early 2000s going to a conference, and putting up a poster saying, “We’ve discovered this phenomenon, but what should we call it?”
There wasn’t a vocabulary – the working vocabulary was largely all based on the Anfinsen principle, and most folks who worked on protein “folding” worked on monomers, not assemblies.
We invented a name for this phenomenon (“morpheeins”), but that was probably a mistake – no one knew the new word, and no one searched for it. I was just the lady in the small lab doing great work with a small group and getting published in high-profile journals. I did not know how to be a mover and shaker. I naively just wanted my colleagues to accept my ideas while I went about the business of running my lab. First they said, “She can’t be right.” Then, “Well this seems to be real for her one protein.” Then, “It seems to be physiologically relevant and a potential drug target for her one protein.” And then, “But surely, this is not a general phenomenon.”
What happened next?
You see where this is going. I started thinking that if my protein does this, and I’ve been working on it for 20 years without realizing it, what other proteins might do this as well?
Turns out there were a number of characteristics that were unusual about this protein, but these unusual phenomena generally didn’t make it into the abstract of a paper. I went looking for those characteristics in other papers. One thing about proteins taking on different assemblies is that it could potentially explain a phenomenon that arose in the 1990s—people called it “protein moonlighting.”
People would use various techniques to find “what protein has this function.” I.e., set up a bacterial system to find a protein that fixes a particular problem. So then the protein would be sequenced, and we’d find out that “hey, we know this protein, it’s a metabolic enzyme, or it works in glycolysis, or is known to have some other very common activity.” Then people realized proteins could do multiple things! Instead of “one sequence, one structure, one function,” they were saying, “One sequence, one structure, multiple functions.” But, in fact, I believe it may well be “one sequence, multiple structures, multiple functions.”
There are now wonderful examples of proteins that take on different assemblies to do different jobs. They are just now being published. My favorite is the enzyme ribonucleotide reductase. It’s twenty years after we saw the octamer turn hexamer and back again and could not figure out what to call the phenomenon. I have no ideas when these ideas will become mainstream.
There are other excellent examples of this now. There are circadian clock proteins, where in order for them to work, they must change their fold. It’s not like a prion—it’s not irreversible, it’s not a disease. These proteins regularly go back and forth between different structures very slowly.
Another example of proteins that can be in different assemblies are those that can form filaments, and then filaments can come apart. Do we understand the function of the protein when it’s a filament? Do we understand the structural differences between proteins when in a filament and when not? In most cases, we do not as yet.
So over the last few years of my career, I have arranged for conference sessions focusing on proteins that do these various forms of shape-shifting. Talks on filaments, on moonlightings, on circadian clock proteins, etc., just to put forward the notion that there are lots of exceptions to “one sequence, one structure, one function,” rule and these exceptions create lots of avenues for designing and discovering drugs. Now we’re talking about drug molecules that might affect this equilibrium of different structures.
This is still largely an unproven idea 20 years later, because hardly anyone is looking for it . . . and though I tried to get a search for such proteins funded, I was not successful.
Say more about the implications for drug development, in your view?
Here’s my hunch: Most drugs fail because they are based on the foundation of “one sequence, one structure, one function.” They’re misguided at the most fundamental level.
Take a step back. The original protein I studied (porphobilinogen synthase) caused an inborn error of metabolism. Fewer than 20 people in the entire world had been diagnosed with this particular disease.
We made these disease-associated proteins, and we found out that in 100% of cases, the equilibrium among the assemblies was unlike the wild type. In other words, this ability to shapeshift was relevant to the disease.
So, what about drug side effects? There are lots of drugs where we don’t understand the mechanism or the mechanism of their side effects. So let’s get hold of a collection of FDA approved drugs, and ask whether any of them have an effect on the assemblies of this protein. A few did shift the equilibrium toward the inactive assembly. So here’s a mechanism for a drug side effect.
Then we asked the question—could we find small molecules that bind to these different structures and either increase or decrease the activity of this protein? By this time, we had the crystal structure of both of assemblies. We used docking to assembly-specific sites to look for small molecules that would stabilize one assembly versus another. Again, success.
Turns out that this protein is in all organisms – it is an enzyme that nearly every cellular organism needs. Plants need this protein to make chlorophyll, and we need it to make hemoglobin. Turns out that there are sequence differences between the plant protein and our version. Indeed, we found a molecule that would inhibit the plant protein but not the human protein. Perhaps a potential herbicide.
Since it was so hard to get our morpheein-related work funded, particularly the search for potential morpheeins, in order to stay in business I decided to look for an inborn error of metabolism that has thousands or tens of thousands of victims, not just 20 people in the world. So let’s look for the protein involved, and let’s ask whether the protein has different assemblies. This caused us to move into the field of phenylketonuria (also known as PKU). It’s tested for by newborn screening, because if it’s not treated early there can be serious brain damage. Before the mid-20th Century, people who had PKU became progressively disabled and generally died young. Now we know one successful treatment is a very rigid low-protein diet. If people with PKU don’t stick to the diet, they end up with behavioral and mental issues, they end up in jail, or other unhappy consequences.
It took several years for us to get funded in the PKU field. We proposed that the PKU protein changed shape in this interesting way. This idea was against the dogma and was met with huge skepticism: “We know the structure of this protein.” We did eventually get that grant funded, about six years ago, but only after we has a PNAS paper on the first crystal structure of the full length mammalian protein. We had to have a big success before we could get our PKU work funded. This is a common problem with getting science funded.
When that grant got funded, I said to myself, “This is great, we’ll work on it for 5 years, hopefully make some good progress, but then I’ve had it with this fight to maintain NIH funding.” It’s been over 40 years. So at this point, I’ve become semi-retired. I’m 68, and this kind of battle to stay funded pushing innovative ideas is for somebody who is younger.
What do you think of AI algorithms that purport to predict protein structure?
I’ve seen some of those predictions, and they are amazing. But they’re predicting a structure for the protein, not necessarily the only structure. So this is still an example of how a dogma has gotten in the way of moving certain ideas forward.
What was your experience with NIH funding?
I submitted applications to NIH (and other agencies) to search for proteins that might be morpheeins. There were many proposals. I submitted for fancy programs, etc. Study sections would read the proposals – 2 to 3 people might read the proposal in detail, and one person reports on it, and everyone else votes based on what these people said. What I would get is: One or two people would think “this is the best thing since sliced bread,” and one or two people would say, “She’s crazy, proteins don’t do that.”
Nearly everything we (as protein scientists) do, and every experiment we design is based on the Anfinsen principle! If we don’t believe “one sequence, one structure, one function,” we’d have to throw out our basic understanding. It’s a new idea (even more so 15 – 20 years ago), and new ideas frighten people.
I ended up entering into an unfunded period of my career. By that time, I was long-tenured, and had decades of continuous NIH support (just not for the new idea).
When I tried to fold the new idea into an existing grant, some study section members went, “Wow, this is great, Eileen is so innovative!,” but others said, “Eileen has gone off the rails.”
But my institution supported people who go unfunded. They continued to support me and my lab at a low level for a few years. In fact, it was our president, Bob Young, who suggested my work for the New York Times article where you first ran across mention of me.
Eventually, I had to let go of the new idea, because if I continued to pursue these ideas doggedly, I’d be retired much earlier than I wanted. My position depended on having a federally-funded lab. So, I left the pursuit of funding for “finding morpheeins”, and turned to PKU, whose central protein, it turns out, might be a morpheein.
One problem with our study section system is that funding (the success rate) is so low that members want to fund studies what will work. If an established member of the community says, “This is a crazy idea,” that grant application will never rise to the top of the pile. And you need to rise to the very top to get funded. Every member of the study section needs to be convinced that your project will work.
What if you had a “high-risk, high-reward program” that instead of asking directly for applications, instead went around looking for grants that got bimodal reviews from any study section?
Yes, that’s a great idea. You should ask the folks who run study sections to pull out those grants where somebody loves it, and somebody else thinks it will never work. Those grants often don’t get discussed at all anymore.
It used to be the case that you could argue in a study section for changing a grant’s rating. I was sitting on a study section in the 1990s. An application was assigned to me, and it involved a timekeeping protein—which we now know can change structure. I argued for it vehemently in study section, but it didn’t get funded. I’m not sure it even scored well.
Back in 1990s, you could argue for something because the payline might have been 25-30 percent. Now that it’s 10-12%, nobody bothers arguing. You’re not going to win, so why waste the effort?
In the past few years, are there any papers you wish you could have published?
I don’t know that I have ever wanted to publish someone else’s paper. However, there have been papers that I find extraordinary, most recently because they do such a spectacular job of supporting the notion that one protein can participate in alternate assemblies, with alternate functions, and alternate interaction partners. One such paper is the review on the alternate forms and functions of ribonucleotide reductase, out of Yimon Aye’s laboratory. It synthesizes an amazing story from the work of her lab and others. Here’s the link: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7131891/.