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Autobiography of Attila Szabo I was born on September 6, 1947, in Budapest, Hungary. My father was a painter, and my mother was a seamstress. Although neither of them went to high school, it was clear from the very beginning that they wanted me to go to university. I was an only child and therefore quite spoiled. My interest in science was aroused by a distant cousin when I was about seven. For reasons that are still unclear to me, he once brought over a bunch of test tubes and flasks filled with liquids of different vivid colors. In addition, he gave my father a red and a white powder. When gently mixed together, rolled in a piece of paper and then stepped on, it exploded violently with a huge bang followed by a tremendous puff of white smoke. This made a lasting impression on me. Since I was born in September, I started school a year older than my friends. When the revolution began in October 1956, I was in the third grade. My parents decided to leave, primarily to give me a better life. My father arranged to get a letter directing him to paint a house near the Austrian border. We got on a train that was filled with people carrying huge suitcases and even Persian carpets. While I don’t know for sure, the Russians probably decided to let all these people escape. If they would have shot a few people on one of those trains, my parents would have never left. Nevertheless, we escaped just like in the movies. We paid some villagers to guide us across the border in the middle of the night. There were even flares being shot up around us (no doubt fired by the villagers to give us our money’s worth), and we had to hit the ground every time. After crossing the border, we were bussed to Vienna. We now had to find a country that would take us. My father decided to go somewhere warm. Painting is a seasonal occupation and he wanted to work the whole year around. All Hungarians wanted to go to the U.S., and consequently, there were huge lines at the embassy. My father did not like to stand in line, so the U.S. was out. He considered Australia, but something went wrong. Argentina seemed like a good choice because they promised us an apartment. Apparently, we came very close to going there. When my father was picking up our plane tickets at the embassy, he overheard two people behind the counter rapidly speaking Spanish. My father said, “Oh, my God, I could never learn this language.” So, Argentina was out. By this time, the only country that would take us was Canada. We were to go to Winnipeg which was far from the warm place we had envisioned. Fortunately, we were put into a holding camp near Montreal. After visiting the city, my father was excited that there were no igloos, and we “escaped” from the camp. This is how we ended up in Montreal in the summer of 1957. Although purely accidental, this turned out to be the ideal choice. I never had to worry about Vietnam. I started school in the fall. I spoke very little English. Because my father was Catholic, I was sent to a public school run by religious brothers. The principal took a look at me and decided that I was just the right size for the fifth grade. So because I was a big kid, I skipped a couple of grades. In addition, high school was only 11 grades at that time. So, all through my education, I was younger than my fellow students, not because I was smart but because I was fat. When I was around 11, my mother bought some shoe polish that came in a bottle shaped like a flask. This sparked something in me, and I got my parents to buy me a chemistry set. I soon 10.1021/jp800825t
got tired of the rather boring experiments (which were presumably exciting compared with those one can do today). So, I bamboozled my father to get me all sorts of dangerous chemicals from our pharmacist. My only real discovery involved lighting a mixture of alcohol and carbon tetrachloride. After breathing the fumes, I felt that my lungs were about to collapse. Perhaps I had found a way of making phosgene. At this time, the major goal of my life was to discover the nature of the two powders that my cousin had given us in Hungary. The central problem facing any young chemist is that it is difficult to get really big bangs. Nitrogen triiodide gives only small pops. One day I was rubbing potassium chlorate crystals against a matchbox to make sparks. I immediately recognized the smell. The two chemicals must have been potassium chlorate and red phosphorus. How could I get my hands on a little red phosphorus? Who would sell some to a 14 year old boy? Well, Fisher Scientific would. I still remember excitedly bicycling home with a large tin of red phosphorus. I could now really impress my friends with ear-deafening explosions followed by enormous puffs of white smoke. At about this time, I started to become interested in theory and, in particular, the nature of chemical bonding. I was soon trying to read some college chemistry textbooks, learning about s, p, d, ... , orbitals and all that. In the spring of 1963, I entered a science fair with a project dealing with atomic structure. I went to McGill in 1964 but continued living with my parents. I took only math, physics, and chemistry classes after my freshman year. The most significant thing I did as an undergraduate was to convince Branka Ladanyi, who was in many of my physics classes, to switch to chemical physics from physics. In 1967, I started thinking about graduate schools and wrote letters to several theoretical chemists including Robert Parr and Martin Karplus. I was admitted to several places but in the end chose Harvard probably because it was the only school my mother had ever heard of. I arrived in Cambridge in September 1968. In the beginning, Harvard was a little intimidating. I had to take a p-chem entrance exam. One of the questions asked for the eigenvalues of a harmonic oscillator with an infinite reflecting wall down the middle. At first I didn’t know how to solve it, and I remember saying to myself that since I wanted to be a quantum mechanic I should be able to do it and it can’t really be all that hard. Eventually, I did figure it out and I was quite proud of myself. After the exam, I overheard two organic chemistry students say, “Oh yeah, that harmonic oscillator problem was really easy!” I had decided to work with Martin. The first problem that he suggested was to analyze NMR experiments on paramagnetic iron-porphyrin complexes done by Kurt Wu¨thrich and Robert Shulman. I had to learn about pseudocontact shifts, Kramers doublets, g tensors, and all that. I worked on this for about a year and my thesis has a long chapter on it, but we never published anything. I did not find this problem too exciting and never imagined I would ever do anything with NMR again. In the spring of 1970, Martin suggested that we go to MIT to hear a series of lectures by Max Perutz on how hemoglobin works. After the lectures were over, Martin told me that the time was right to think about making a model for hemoglobin.
This article not subject to U.S. Copyright. Published 2008 by the American Chemical Society Published on Web 05/08/2008
5884 J. Phys. Chem. B, Vol. 112, No. 19, 2008 I seem to remember thinking why in the world would he want me to make a ball and stick model of the structure of hemoglobin! Fortunately, it soon became clear that what Martin had in mind was a statistical mechanical model. He wanted me to construct a model of Perutz’s mechanism that could be used to quantatively understand the vast amount of experimental data available in the literature. Coincidentally, the year before, I went to a lecture by Harden McConnell where he gave the formulas for the binding curves obtained by Linus Pauling for some simple models of cooperativity. At the same time, I was taking a course from Roy Gordon who just showed us how easy it is to treat a lattice gas using the grand canonical partition function. Just for fun, I rederived Pauling’s results using this technique. So, a year later, I could really impress Martin by tackling the problem using this elegant formalism. If it were not for this happy coincidence, I am sure that using the grand partition function would have never occurred to me. I find it remarkable that my approach to science was so different then. I simply had too much energy. I began by reading Perutz’s two famous 1970 Nature papers countless times and then proceeded to formulate the most faithful representation of his ideas including all the bells and whistles. I considered both oxygen and proton binding and several different kinds of “saltbridges”. Needless to say, the partition function was a sum of many terms and looked pretty complicated. Then, one night, as we were writing all this up, I started fooling around by combining various terms. To my amazement, I soon discovered that the entire partition function could be written in a simple form that had the same structure as that of the MonodWyman-Changeux allosteric model. So, in effect, what we did was provide a microscopic, structural interpretation of their phenomenological parameters. Had I started by formulating the simplest possible model that captures the essence of Perutz’s mechanism, I would have seen this immediately. My experience in graduate school was truly wonderful. I learned so much from so many people. At that time, all theoretical grad students and postdocs had their offices in an old detached home called Prince House. A remarkable number of now well-known theorists were there at one time or another. They worked on a wide variety of topics and were more than willing to talk about their work to anyone who would listen. I am a good listener. I do not wish to mention them all by name because I am afraid I’ll leave someone out. So, I will focus on two of my office mates who had the biggest influence on my scientific life. Neil Ostlund was Martin’s postdoc and a real quantum chemist, being a student of John Pople. Since quantum chemistry was my first love, I spent a lot of time talking to him and others in Prince House who were applying many-body theory to electronic structure problems. Neil showed me how one can understand the relationship between different sophisticated approaches by applying them to a simple minimal basis model of the hydrogen molecule. A few years later, when he was at Arkansas and I was in Indiana, we decided to write a small book that would make complicated things look easy. We worked on this with great enthusiasm all through the late 1970s, and the scope of the book expanded. We each gave what was the best in us, fighting over every word and equation. It was a true collaboration. We are still very happy that we were crazy enough to have written a book so early in our careers and that many people seem to have actually learned something from it. The other person was Klaus Schulten. He arrived in the fall of 1969 and ended up working with both Roy Gordon and Martin Karplus. We became good friends and saw many sunrises
together from our office window. His impact on my science really started when in 1976 he and Zan Luthey-Schulten invited me to spend the summer at the Max Planck Institute for Biophysical Chemistry in Go¨ttingen, but more about this later. In the fall of 1971, I found out that Martin was planning to go on sabbatical in Paris the next year. So, I modestly suggested to him that if he would give me my Ph.D. after just four years in grad school, I could go to France with him as a postdoc and it would not cost him a penny. I was Canadian, and pretty sure that I could get a fellowship from the National Research Council. Happily, Martin went along with this and I arrived in Paris in September 1972 to continue working with him on hemoglobin. The next year, I went to Cambridge, England. Since I had my own money, I had desks both in the Department of Theoretical Chemistry with David Buckingham and at the Medical Research Council Laboratory of Molecular Biology with Max Perutz. I began worrying about trying to find a job. I thought I would go back to Canada, but I ended up accepting an offer from Indiana, where Peter Langhoff, a former Karplus postdoc, had a faculty position. I arrived in Bloomington in August 1974 and started to worry about what kind of research to do. I will now try to describe in some detail how I ended up working on different problems that became life-long interests. Hopefully some readers who are just starting their careers will find it interesting. In the course of my work on using linear free energy relations to try to understand hemoglobin kinetics, I wondered what the maximum rate of ligand binding would be. Thus began my longterm love affair with diffusion-controlled reactions. At first, I worked on extending Smoluchowski’s calculation of the rate constant to anisotropic (e.g., a small absorbing hole on the surface of a protein) and fluctuating (e.g., the hole opens and closes) reactivity. Then, one day I realized how little I understood. On the molecular level, an irreversible bimolecular reaction can occur only once. So, what does this have to do with the steady-state flux to an absorbing surface that can repeatedly kill particles? Although this puzzle was solved long ago, the answer is still not widely appreciated. In any case, I started to think about the many-particle aspects of irreversible and then reversible bimolecular reactions. It turned out to be challenging to develop a microscopic theory of the kinetics of reversible diffusion-influenced reactions on all time scales. One reason is that the relaxation to equilibrium does not follow an exponential time course but rather a power law. I started working on this in the late 1980s with Noam Agmon. It is only relatively recently that Irina Gopich and I managed to formulate a general theory that has a remarkably simple structure. One day, I was talking to another assistant professor, Mark Wightman, who was sticking microelectrodes into rat brains to measure neurotransmitter levels. His electrodes were made by embedding an ultrathin carbon fiber into a glass rod. It occurred to me that the problem of calculating the current to such an electrode is essentially the same as obtaining the flux through a small hole in a protein. So, I actually knew something about his problem without realizing it. Because one of my high school science fair projects dealt with the electrolysis of water, I thought it would be fun to do something in electrochemistry, and I ended up writing about ten papers in this area. In the summer of 1976, I went to visit Klaus and Zan Schulten in Go¨ttingen. I stayed in a guest house at the Institute, and since I do not speak a word of German, all I could do was work (and eat and drink). Klaus suggested that we work on something completely different. One of his colleagues had started doing experiments on exciplex formation of two optical probes
J. Phys. Chem. B, Vol. 112, No. 19, 2008 5885 attached to the ends of a polymer, so Klaus proposed we work out the theory. I had no idea how to start. I knew about Paul Flory’s book on polymer chemistry but had only looked at it, and it didn’t seem to discuss kinetics. As I later found out, Marshall Fixman wrote a couple of important papers on this subject around this time. If I had seen them, I would have probably stopped immediately because they looked so complicated. So, sometimes ignorance is not really such a bad thing. I began by making up the simplest possible model that I had some chance of solving. Needless to say, it had to be onedimensional. My polymer was made up of unit vectors or bonds that could independently point either left or right. Each bond could flip with a certain rate. Amusingly, this model was the same as the one Bob Zwanzig, Biman Bagchi, and I used many years later to illustrate Levinthal’s “paradox” in protein folding. The ends of this polymer touch when equal numbers of bonds point left or right, and we wanted to know the mean time it takes to reach this conformation. It was easy to write down the set of discrete rate equations for the end-to-end distance distribution because they are the same as those that describe the kinetics of ligand binding to a “super-hemoglobin” with N noninteracting sites. Late one night, it occurred to me that one could make life simpler by taking the continuum limit. In this way, I derived a partial differential equation for the probability distribution of the end-to-end distance. Although I was too ignorant to know at the time, it turned out to be simply the Smoluchowski equation for diffusion on a harmonic surface. All of this is so obvious to me now that it is hard to believe how much I struggled then. So, to find the distribution of lifetimes of our one-dimensional polymer with reactive ends, we had to solve this equation subject to an absorbing boundary condition at the minimum of the potential. I immediately realized that this problem is almost exactly the same as the one on my Harvard entrance exam that I mentioned previously. Using the properties of Hermite polynomials, I expressed the survival probability analytically as an infinite series. At the same time, Klaus was also trying to solve this problem. He was aware of the analytic expression for the Green’s function of a diffusive oscillator and then used the method of images to get an extremely simple expression for the survival probability (the inverse sine of an exponential). I am still jealous. We then looked up the Taylor series expansion of the inverse-sine, and lo and behold, the two expressions were identical. It’s fun to collaborate. We generalized our model to three dimensions by describing the stochastic dynamics of the polymer as diffusion under the influence of the potential of mean force between the reactive ends. This is the simplest model that describes relaxation to the equilibrium distribution of end-to-end distances. Our excitement soon faded because we could not make any progress in analytically calculating the kinetics. On my next trip to Go¨ttingen, we tried again. One day, I was looking at a newly arrived book in the library, called “Stochastic Models in Biology”, by Goel and Richter-Dyn. It seemed pretty complicated to me, but somehow, I managed to see that it contained the answer to all our prayers. It showed that, if one is interested only in the mean lifetime (which is the same as the mean first passage time to an absorbing boundary), one can bypass the solution of the time dependent diffusion equation and only solve a simple inhomogeneous differential equation. All the rest was easy, but it took us a couple of years to actually write it up. Zan Schulten had developed an efficient finite-difference algorithm for solving the Smoluckowski equa-
tion which she used to test a single-exponential approximation to the survival probability we proposed. In addition, she made an important theoretical contribution to the paper. I generalized the formalism to handle partially absorbing boundaries but I used the free-diffusion form of the boundary condition even in the presence of a potential. This seems manifestly wrong. However, Zan was able to prove that in the context of first passage times it was actually correct. So, it does not hurt to be a little lucky sometimes. Our paper lay dormant for a long time. It eventually did attract the interest of experimentalists, mostly in the area of protein folding, which started when Bill Eaton walked in my office one day and asked if I knew how the rate of loop formation scaled with length. In the 1970s, Indiana was a hotbed of NMR because of Adam Allerhand, a pioneer in the use of wide-bore magnets and an expert in 13C NMR relaxation. He got Frank Gurd, a biochemist, interested in using NMR to study the dynamics of side chains in myoglobin. Frank had a graduate student, Dick Wittebort, who one day came to my office to ask me for help with some papers he was trying to understand. I told him that I knew nothing about NMR relaxation but I did know something about spherical harmonics and Wigner rotation matrices because of all the quantum mechanics courses I had taken. In this modest way, we began a collaboration that had perhaps the greatest impact on my scientific life. We learned lots of new things together. Although I had heard of correlation functions before, this was the time when I actually learned how to calculate them. We then began to make existing models more realistic, for example, by considering restricted rotational diffusion about carbon-carbon bonds in amino acid side chains. Alternatively, to treat excluded volume, we described side-chain dynamics as jumps between discrete configurations on a diamond lattice. We worked on this for a couple of years, and in 1978, we published our first (and my most complicated) paper on NMR. Then, Dick graduated and went to MIT as a postdoc. A new graduate student from Italy, Giovanni Lipari, arrived and for some reason, wanted to work with me. I suggested that he continue where Dick left off and think about the dynamics of DNA. At that time, I went to a seminar on fluorescence depolarization of probes embedded in liquid crystals. This was another topic I knew nothing about, but as I listened, it became increasingly evident that the underlying theory was closely related to that used to describe NMR. This probably was the first time I ever heard the word “order parameter”. Trying to understand fluorescence forced me to think in the time domain, and this makes life easier than thinking in the frequency domain (i.e., spectral densities and all that). Giovanni found that he could analyze experimental data equally well using a variety of physically different models of internal dynamics. Moreover, the parameters of more complex and hence more physically realistic models could not be uniquely determined. This made us reexamine the underlying theory, and it was soon clear that the measured relaxation parameters simply do not depend on all the details of the internal dynamics when the dynamics are sufficiently fast so as to be in the so-called extreme narrowing limit. In this limit, the NMR experiment cannot tell you if a bond is rotating about an axis or wobbling in a cone when the corresponding order parameters are the same. This is why we called our approach to the analysis of NMR relaxation experiments “model-free”. We knew all of this before coming to the NIH, but it took us a couple of years to write it up because we wanted to do the best job possible. Giovanni died in an accident before the proofs arrived, a tragedy that absolutely devastated everyone who knew
5886 J. Phys. Chem. B, Vol. 112, No. 19, 2008 him. When he first came to Indiana, he told me that his mentor in Italy advised him not to work with an unknown assistant professor. In general, this is sound advice and something I can see myself saying. Over the years, I have thought many times that he should have taken it. He most probably would still be alive if he had followed a different timeline. I managed to work on NMR relaxation without ever learning Redfield theory or even reading the relevant chapter in Abragam. However, for some reason that escapes me now, I carefully studied the section in Abragam on Anderson’s theory of line shapes when the frequency fluctuates because of chemical exchange. This was perhaps the most influential single thing I ever read. It is pretty obvious that one can use the same formalism (by making the frequency imaginary) to describe irreversible unimolecular kinetics with fluctuating rates. It is more surprising that it would end up playing a crucial role in my recent work, not only with Irina Gopich on analyzing singlemolecule fluorescence experiments but also with Gerhard Hummer on extracting free energy profiles from non-equilibrium pulling experiments. Ultimately, the reason that all of these physically different problems are mathematically so closely related is that, in the absence of fluctuations, the magnetization in NMR, the survival probability in kinetics, the generating function for Poissonian photon statistics, and the Boltzmann factor are all single exponentials. In the fall of 1979, I came up for tenure. For a while, there were all kinds of rumors why I did not get it. The reason was simply that the chairman felt that, while I was a good scientist, I was not the kind of person who would catapult the department to the next level. This was not such an unreasonable assessment at that time. I was a bit unfocused, working on NMR, atomic physics, hemoglobin kinetics, and finishing a book on quantum chemistry (an area in which I wrote only a handful of papers). Although I never told my parents, I was never too upset about all this, even though I tried to cause as much trouble as possible by exhausting all appeals. I suppose I never really liked living in a small town like Bloomington and thought that this was just an opportunity to get away from there. Not getting tenure at Indiana is a little bit different than not getting tenure at Harvard. My options seemed rather limited even though I just had my NIH grant renewed. Then, I got lucky.
Martin Karplus told Bill Eaton about my predicament and said that I was smart. This was basically it. Bill created a job for me, and I arrived at the NIH in December of 1980. The NIH turned out to be the perfect place for me. I have been given complete freedom to do whatever I want. I am surrounded by great and highly interactive scientists. I do not have to apply for grants, which I always hated because I never know what I am going to do next. Since I do not have to teach or write grants, I have plenty of time to be intimately involved with all the details of the research I am doing. As you can see from my bibliography, I had the privilege to work with many people, both theorists and experimentalists. I would like to take this opportunity to thank all of my collaborators for making my scientific life both fun and reasonably productive. I like to think that the Laboratory of Chemical Physics at the NIH is the best biophysical chemistry lab in the world. It was started in the 1940s by F. S. Brackett of hydrogen atom fame. It was built up by Bill Eaton based on a simple idea: Hire the best scientists you can, and do not worry too much about what they are working on at the moment. It has been a great place to work. The lab has some reasonably well-known NMR spectroscopists: Ad Bax, Marius Clore, and Rob Tycko. The first two are among the top 20 most-cited chemists in the world. Our higher-frequency spectroscopists are not too bad either: Phil Anfinrud, Bill Eaton, Jim Hofrichter, and Ira Levin. Ira, who pioneered two-dimensional infrared imaging, has been a major force in maintaining the tradition of strong support for basic science in our institute. Finally, there are the theoreticians Gerhard Hummer, Eric Henry, Irina Gopich, Bob Zwanzig, and our most recent addition, Artur Adib (Bob, who joined us in the late 1980s, just retired but still comes to the Lab as a Scientist Emeritus). I have learned a great deal from all of them. I cannot imagine having nicer or more stimulating colleagues. Although I have some good stories about the 27 years I spent at the NIH, I have run out of steam. I realize that it is unusual not to spend more time talking about the most important part of one’s career, but in this way, I can at least avoid boring you even further. Attila Szabo
