A preliminary report on my life in science.

I describe my wanderings from the United States to East Germany and back. I hope this gives a glimpse of science in East Germany and encourages people who do science under less than favorable conditions. Although elements of my story are unique, the main points are general: don't be afraid to start something new; it pays to be persistent; and science is a passion--if it feels like fun, you've probably got it right.

ESSAY

I hesitate to write an article about my career. I am told, however, that people may find my story interesting and my experiences of value. It is also an opportunity to express my views on science.

My life has been heavily influenced by the turmoil of the past century. My father was born in Russia into a Jewish family that emigrated during the Russian revolution to Vienna. He was strongly influenced by the social aspirations and political atmosphere of the 1920s and became a member of the Socialist party and later of the Communist party. He studied medicine and chemistry. While he was on a fellowship in Cincinnati, OH, the Nazis took over Austria and he could not return. In Cincinnati, he met my half-Jewish mother, who had emigrated from Nazi Germany and restarted her career as a pediatrician. I was born in Cincinnati, but 3 years later, in 1950, my parents became targets of the anticommunist campaign of Joseph McCarthy and returned to Europe, initially for a year in Austria. Because my dad was blacklisted, he could not find a job in Austria and we moved to East Germany, where he joined the faculty of Humboldt-University in East Berlin. I was 4 years old when we arrived and I stayed in Berlin until the age of 48, when, in 1995, I moved back to the United States, taking a position at the Harvard Medical School in Boston. This turned out to be “scientific heaven,” and I am eternally grateful to Marc Kirschner, who recruited me. I was also fortunate to become a Howard Hughes Medical Institute investigator only 2 years later.

Tom A. Rapoport

My parents are responsible for my early interest in science, often by stimulating discussions at the dinner table. I was first attracted to mathematics, largely because I loved the thought-problems posed in Math Olympiads. I was successful enough in these competitions to enter a special high school for math and science. I soon realized that the problems in Math Olympiads were not actual mathematics (as my brother, who became a real mathematician, always enjoyed pointing out to me), and I discovered my love for chemistry, stimulated largely by Pauling's General Chemistry. I started studying chemistry, but after 3 years I switched to biochemistry. This was somewhat awkward, because my dad was the head of the Biochemistry Institute. That was not an advantage; for example, I once caused a flood and my dad punished me by making me repaint the damaged rooms, including his office.

East Germany had just established an unusual mechanism to obtain a Ph.D.: you could team up with someone else. So, I actually obtained my degree together with a friend Wolfgang Hoehne. We worked in enzymology, on the mechanism of inorganic pyrophosphatase.

Toward the end of my Ph.D., I met Reinhart Heinrich, a theoretical physicist whom my dad hired to do mathematical modeling of metabolism. We hit it off immediately. Prodded by Dad, we developed a concept that describes in quantitative terms the importance of an enzyme for the overall flux through a metabolic pathway and for the regulation of metabolite concentrations (Heinrich and Rapoport, 1973). The theory is now known as “metabolic control analysis (MCA)” and was independently developed by Kascer and Burns (1973). The mathematical modeling work led Reinhart and myself in 1979 to a joint Habilitation or Doktor B, the second degree that you need in Germany for an eventual professorship. Again, I did the thesis in a team! Our main publication is still my most quoted paper (Heinrich and Rapoport, 1974). Indeed, MCA is one of the first contributions to “systems biology.” Over the years, I repeatedly collaborated with Reinhart, often on projects originating from my experimental work. Unfortunately, after 36 years of collaboration and deep friendship, he died suddenly in 2006 at the age of 60.

In parallel with the theoretical work, I also entered molecular biology. I joined the group of Sinaida Rosenthal at the Institute of Molecular Biology of the Academy of Sciences in Berlin-Buch. Sina wanted to establish gene technology in the country and decided to clone insulin mRNA. I was given the task of purifying the mRNA but soon realized that human insulin was beyond our reach. However, I found out that the islets of Langerhans in fish (Brockmann bodies) are much bigger than in mammals and that they are separated from the exocrine pancreas. I decided to use carp, a popular fish that is raised mostly for the New Year's celebration in Germany. On fish days, the entire department would help us. We would stand on both sides of a long table, the cleaning lady would kill the carp, and we would remove the Brockmann bodies. The rest was sold cheap to people lining up in front of the lab, which made us very popular in Berlin. It took us many years to actually clone carp insulin, but we finally managed, several years after Gilbert published on human insulin. Along the way, we determined the first protein and gene sequences in East Germany.

Nothing was easy in East Germany. Most chemicals and materials had to be purchased from the West, and the budget for my group was only ∼US$1000 per year. In addition, we had to place orders a year in advance. We therefore set up a system where each institute would make some chemicals and we would trade them (my job was to make 100 mCi of [35S]methionine every few months). We had to wash Eppendorf tubes and pipette tips, and journals arrived months late. Contacts with and travel to the West were restricted. I was lucky to be permitted to travel to the United States (after 10 years). I cannot thank enough those American scientists who helped me during this time (Don Steiner, Harvey Lodish, Gunter Blobel, Peter Walter, Lila Gierasch, Rick Klausner, and others). I would usually give seminars at several places, and use the honoraria to purchase chemicals and materials. The FBI would call each host after my visit to find out whether I was a “real scientist.” On the way back, I would often travel with so many packages (ice and dry ice included) that, at the airport, my packages and I would move like an inchworm; I would advance some, go back and bring up the rear, and move forward again. Luckily, airport security was not as strict as today.

Despite all these obstacles, we did manage to do science and have fun with it. Because of our budget restrictions, we had to plan every experiment in detail. We would even discuss the order in which samples would be loaded onto a gel. Obviously, this forced us to think carefully about the controls and possible conclusions—great training for everyone in the lab. From my own experience, I can say that it is possible to contribute to the progress of science even when working under unfavorable conditions. The most important thing is to generate an exciting atmosphere in the lab, in which informal discussions happen at the bench; papers are trashed in an irreverent manner; ideas are tossed around; and experiments are suggested, even if you cannot perform them. This can be done anywhere. Biological science is a communal effort, despite the impression that you sometimes get from prizes. Being part of this international family of scientists is one of the best aspects of our job.

Cloning carp insulin mRNA brought me into the field of protein translocation. Translation of the mRNA in vitro produced preproinsulin, which I realized after hearing Gunter Blobel present his “signal hypothesis.” I became interested in how the signal sequence is recognized and how a polypeptide moves through the membrane. Our first real success came in collaboration with the late Sasha Girshovich and his wife, Lena Bochkareva, then at the Institute for Protein Research in Poustchino (Russia). Based on the demonstration by Art Johnson that ε-acetylated lysyl-tRNA is incorporated into polypeptides (Johnson ), we attached a photoreactive group to the amino acid and showed that the signal sequence of a nascent polypeptide can be cross-linked to a subunit of the signal recognition particle (Kurzchalia ). We later used photocross-linking to demonstrate that Sec61p, discovered in a genetic screen by the Schekman lab, surrounds the polypeptide chain during its passage through the membrane (Mothes ), providing the first evidence that Sec61p forms the protein-conducting channel. This was strongly supported by reconstitution experiments with purified translocation components, carried out by my student Dirk Görlich, who demonstrated that the Sec61p complex, a heterotrimeric membrane protein complex consisting of Sec61p and two small polypeptides, is the essential membrane component (Görlich and Rapoport, 1993). We also realized that the eukaryotic Sec61 and the bacterial SecY complexes are related, and this together with reconstitution experiments of Escherichia coli components by the Wickner and Mizushima labs led to a unifying concept of protein translocation. This was an exciting time in the lab, and our success was facilitated by our sudden “wealth” after unification of Germany. In fact, when I left Germany, I had seven grants, all obtained at the requested budget. At one point, we used our funds to purchase the world's supply of digitonin! But, I also had a great team of students, many of whom, I am proud to say, are now professors.

When I moved to the United States in 1995, most of my students came with me, all on the same airplane. They were soon known throughout the department as “The Germans.” We continued to make important discoveries on the mechanism of protein translocation, but I soon realized that any real breakthrough required structural information about the channel. We started with electron microscopy in collaboration with Chris Akey at Boston University but then decided to determine an x-ray structure of the SecY complex. Most people thought we were crazy, and, frankly, I was not very optimistic myself. However, in collaboration with Steve Harrison, we finally succeeded (Van den Berg ). Many factors contributed, including choosing the right detergent, using an archaebacterium as the source of SecY, and expressing all three subunits of the complex. Much of the credit goes to three postdocs in the lab: Bert van den Berg, Bil Clemons, and Ian Collinson. It was incredibly exciting to see the structure of the channel slowly emerge as we improved the model and it all began to make sense. We could explain how the channel opens, how it integrates membrane proteins, and how it maintains the barrier for small molecules when translocating a polypeptide. It was an adventure learning x-ray crystallography, and although I cannot claim to be a real structural biologist, I hope others feel encouraged to make a similarly bold move. More recently, we have determined the structure of a complex consisting of SecY and the SecA ATPase (Zimmer ), which clarified how SecA pushes polypeptides through the channel. After more than 30 years in protein translocation, I can say that persistence has paid off. Yet, so many interesting and challenging questions remain to be answered. One of our major goals for the future is to get the structure of the channel in action, i.e., with a translocating polypeptide.

Soon after I moved to Boston, my lab started many entirely new projects. Over the years, we have worked on kinesin, on the movement of cholera toxin and of nonenveloped viruses into the cytosol, on the mechanism of the chaperone BiP, on the transport of DNA across membranes during sporulation of Bacillus subtilis, and on the structure and mechanism of vitamin K epoxide reductase. Most of these projects left the lab with the people doing them. However, two became “permanent” interests of mine. One project concerns the mechanism by which misfolded proteins move from the endoplasmic reticulum (ER) into the cytosol, where they are degraded by the proteasome, a pathway called endoplasmic reticulum–associated protein degradation (ERAD). Over the years, we have identified several ERAD components, but how proteins cross the membrane remains mysterious. The other major project in the lab concerns how the ER achieves its shape. Starting with an in vitro assay that recapitulates the formation of an ER network in a test tube, we have identified membrane proteins that are required for the formation of ER tubules and GTPases that fuse these tubules to form a network. More recently, we became interested in how ER sheets are generated. We started this new field almost a decade ago, and there remains much to be learned.

I am always surprised when young people tell me that they feel that everything important has already been discovered and nothing is left for them. In my experience, wherever you dig, you find gold. Sometimes you set out to solve a particular problem, but other times you just stumble over it. I feel that you should always address an important and interesting biological question, because life is too short to waste on minor details. But, you have to realize that it may take a long time to get somewhere, usually at least 10 years. Training in different areas is extremely helpful, because it facilitates transitions into new fields and gives confidence that you can make even radical changes.

I am a strong advocate for research that is aimed at understanding the mechanism of a biological process. For me, the ideal project begins with a complex phenomenon in an intact cell and leads all the way to a defined system, in which the process is recapitulated with purified molecules and understood at a structural and mechanistic level. Of course, there are other ways to do science, but I feel that we currently see too much emphasis on the latest technological fashion, often with rather descriptive or superficial results. There is nothing that can compare with the excitement of a real insight and discovery. For me, a nice experiment, however small, can make my day. And, I love to propose bold, unifying models, much to the horror of my young coworkers, who are concerned that they may turn out to be wrong (and often are!). It is wonderful to have a democratic, collegial, and humorous atmosphere in the lab; to share the excitement with students and postdocs; and to see them develop into independent investigators.