Though this transformation phenomenon seemed to defy all understanding, Griffith's observations at first created little stir in the scientific world. This was partly because Griffith was intensely private and so averse to large gatherings that he seldom attended scientific conferences. Once, he had to be virtually forced to give a lecture. Bundled into a taxi and escorted to the hall by colleagues, he discoursed in a mumbled monotone, emphasizing an obscure corner of his microbiological work but making no mention of bacterial transformation. Luckily, however, not everyone overlooked Griffith's breakthrough.
Oswald Avery was also interested in the sugarlike coats of the Pneumococcus. He set out to duplicate Griffith's experiment in order to isolate and characterize whatever it was that had caused those R cells to change to the S type. In 1944 Avery, MacLeod, and McCarty published their results: an exquisite set of experiments showing unequivocally that DNA was the transforming principle. Culturing the bacteria in the test tube rather than in mice made it much easier to search for the chemical identity of the transforming factor in the heat-killed S cells. Methodically destroying one by one the biochemical components of the heat-treated S cells, Avery and his group looked to see whether transformation was prevented. First they degraded the sugarlike coat of the S bacteria. Transformation still occurred: the coat was not the transforming principle. Next they used a mixture of two protein-destroying enzymes, trypsin and chymotrypsin, to degrade virtually all the proteins in the S cells. To their surprise, transformation was again unaffected. Next they tried an enzyme (RNase) that breaks down RNA (ribonucleic acid), a second class of nucleic acids similar to DNA and possibly involved in protein synthesis. Again transformation occurred. Finally, they came to DNA, exposing the S bacterial extracts to the DNA-destroying enzyme, DNase. This time they hit a home run. All S-inducing activity ceased completely. The transforming factor was DNA.
In part because of its bombshell implications, the resulting February 1944 paper by Avery, MacLeod, and McCarty met with a mixed response. Many geneticists accepted their conclusions. After all, DNA was found on every chromosome; why shouldn't it be the genetic material? By contrast, however, most biochemists expressed doubt that DNA was a complex enough molecule to act as the repository of such a vast quantity of biological information. They continued to believe that proteins, the other component of chromosomes, would prove to be the hereditary substance. In principle, as the biochemists rightly noted, it would be much easier to encode a vast body of complex information using the twenty-letter amino-acid alphabet of proteins than the four-letter nucleotide alphabet of DNA. Particularly vitriolic in his rejection of DNA as the genetic substance was Avery's own colleague at the Rockefeller Institute, the protein chemist Alfred Mirsky. By then, however, Avery was no longer scientifically active. The Rockefeller Institute had mandatorily retired him at age sixty-five.
Avery missed out on more than the opportunity to defend his work against the attacks of his colleagues: He was never awarded the Nobel Prize, which was certainly his due, for identifying DNA as the transforming principle. Because the Nobel committee makes its records public fifty years following each award, we now know that Avery's candidacy was blocked by the Swedish physical chemist Einar Hammarsten. Though Hammarsten's reputation was based largely on his having produced DNA samples of unprecedented high quality, he still believed genes to be an undiscovered class of proteins. In fact, even after the double helix was found, Hammarsten continued to insist that Avery should not receive the prize until after the mechanism of DNA transformation had been completely worked out. Avery died in 1955; had he lived only a few more years, he would almost certainly have gotten the prize.
When I arrived at Indiana University in the fall of 1947 with plans to pursue the gene for my Ph.D thesis, Avery's paper came up over and over in conversations. By then, no one doubted the reproducibility of his results, and more recent work coming out of the Rockefeller Institute made it all the less likely that proteins would prove to be the genetic actors in bacterial transformation. DNA had at last become an important objective for chemists setting their sights on the next breakthrough. In Cambridge, England, the canny Scottish chemist Alexander Todd rose to the challenge of identifying the chemical bonds that linked together nucleotides in DNA. By early 1951, his lab had proved that these links were always the same, such that the backbone of the DNA molecule was very regular. During the same period, the Austrian-born refugee Erwin Chargaff, at the College of Physicians and Surgeons of Columbia University, used the new technique of paper chromatography to measure the relative amounts of the four DNA bases in DNA samples extracted from a variety of vertebrates and bacteria. While some species had DNA in which adenine and thymine predominated, others had DNA with more guanine and cytosine. The possibility thus presented itself that no two DNA molecules had the same composition.
At Indiana I joined a small group of visionary scientists, mostly physicists and chemists, studying the reproductive process of the viruses that attack bacteria (bacteriophages – "phages" for short). The Phage Group was born when my Ph.D. supervisor, the Italian-trained medic Salvador Luria and his close friend, the German-born theoretical physicist Max Delbrück, teamed up with the American physical chemist Alfred Hershey. During World War II both Luria and Delbrück were considered enemy aliens, and thus ineligible to serve in the war effort of American science, even though Luria, a Jew, had been forced to leave France for New York City and Delbrück had fled Germany as an objector to Nazism. Thus excluded, they continued to work in their respective university labs – Luria at Indiana and Delbrück at Vanderbilt – and collaborated on phage experiments during successive summers at Cold Spring Harbor. In 1943, they joined forces with the brilliant but taciturn Hershey, then doing phage research of his own at Washington University in St. Louis.
The Phage Group's program was based on its belief that phages, like all viruses, were in effect naked genes. This concept had first been proposed in 1922 by the imaginative American geneticist Herman J. Muller, who three years later demonstrated that X rays cause mutations. His belated Nobel Prize came in 1946, just after he joined the faculty of Indiana University. It was his presence, in fact, that led me to Indiana. Having started his career under T. H. Morgan, Muller knew better than anyone else how genetics had evolved during the first half of the twentieth century, and I was enthralled by his lectures during my first term. His work on fruit flies (Drosophila), however, seemed to me to belong more to the past than to the future, and I only briefly considered doing thesis research under his supervision. I opted instead for Luria's phages, an even speedier experimental subject than Drosophila: genetic crosses of phages done one day could be analyzed the next.
For my Ph.D. thesis research, Luria had me follow in his footsteps by studying how X rays killed phage particles. Initially I had hoped to show that viral death was caused by damage to phage DNA. Reluctantly, however, I eventually had to concede that my experimental approach could never give unambiguous answers at the chemical level. I could draw only biological conclusions. Even though phages were indeed effectively naked genes, I realized that the deep answers the Phage Group was seeking could be arrived at only through advanced chemistry. DNA somehow had to transcend its status as an acronym; it had to be understood as a molecular structure in all its chemical detail.
Upon finishing my thesis, I saw no alternative but to move to a lab where I could study DNA chemistry. Unfortunately, however, knowing almost no pure chemistry, I would have been out of my depth in any lab attempting difficult experiments in organic or physical chemistry. I therefore took a postdoctoral fellowship in the Copenhagen lab of the biochemist Herman Kalckar in the fall of 1950. He was studying the synthesis of the small molecules that make up DNA, but I figured out quickly that his biochemical approach would never lead to an understanding of the essence of the gene. Every day spent in his lab would be one more day's delay in learning how DNA carried genetic information.
My Copenhagen year nonetheless ended productively. To es
cape the cold Danish spring, I went to the Zoological Station at Naples during April and May. During my last week there, I attended a small conference on X-ray diffraction methods for determining the 3-D structure of molecules. X-ray diffraction is a way of studying the atomic structure of any molecule that can be crystallized. The crystal is bombarded with X rays, which bounce off its atoms and are scattered. The scatter pattern gives information about the structure of the molecule but, taken alone, is not enough to solve the structure. The additional information needed is the "phase assignment," which deals with the wave properties of the molecule. Solving the phase problem was not easy, and at that time only the most audacious scientists were willing to take it on. Most of the successes of the diffraction method had been achieved with relatively simple molecules.
My expectations for the conference were low. I believed that a three-dimensional understanding of protein structure, or for that matter of DNA, was more than a decade away. Disappointing earlier X-ray photos suggested that DNA was particularly unlikely to yield up its secrets via the X-ray approach. These results were not surprising since the exact sequences of DNA were expected to differ from one individual molecule to another. The resulting irregularity of surface configurations would understandably prevent the long thin DNA chains from lying neatly side by side in the regular repeating patterns required for X-ray analysis to be successful.
It was therefore a surprise and a delight to hear the last-minute talk on DNA by a thirty-four-year-old Englishman named Maurice Wilkins from the Biophysics Lab of King's College, London. Wilkins was a physicist who during the war had worked on the Manhattan Project. For him, as for many of the other scientists involved, the actual deployment of the bomb on Hiroshima and Nagasaki, supposedly the culmination of all their work, was profoundly disillusioning. He considered forsaking science altogether to become a painter in Paris, but biology intervened. He too had read Schrödinger's book, and was now tackling DNA with X-ray diffraction.
He displayed a photograph of an X-ray diffraction pattern he had recently obtained, and its many precise reflections indicated a highly regular crystalline packing. DNA, one had to conclude, must have a regular structure, the elucidation of which might well reveal the nature of the gene. Instantly I saw myself moving to London to help Wilkins find the structure. My attempts to converse with him after his talk, however, went nowhere. All I got for my efforts was a declaration of his conviction that much hard work lay ahead.
While I was hitting consecutive dead ends, back in America the world's preeminent chemist, Caltech's Linus Pauling, announced a major triumph: he had found the exact arrangement in which chains of amino acids (called polypeptides) fold up in proteins, and called his structure the a-helix (alpha helix). That it was Pauling who made this breakthrough was no surprise: he was a scientific superstar. His book The Nature of the Chemical Bond essentially laid the foundation of modern chemistry, and, for chemists of the day, it was the Bible. Pauling had been a precocious child. When he was nine, his father, a druggist in Oregon, wrote to the Oregonian newspaper requesting suggestions of reading matter for his bookish son, adding that he had already read the Bible and Darwin's Origin of Species. But the early death of Pauling's father, which brought the family to financial ruin, makes it remarkable that the promising young man managed to get an education at all.
As soon as I returned to Copenhagen I read about Pauling's a-helix. To my surprise, his model was not based on a deductive leap from experimental X-ray diffraction data. Instead, it was Pauling's long experience as a structural chemist that had emboldened him to infer which type of helical fold would be most compatible with the underlying chemical features of the polypeptide chain. Pauling made scale models of the different parts of the protein molecule, working out plausible schemes in three dimensions. He had reduced the problem to a kind of three-dimensional jigsaw puzzle in a way that was simple yet brilliant.
Whether the a-helix was correct – in addition to being pretty – was now the question. Only a week later, I got the answer. Sir Lawrence Bragg, the English inventor of X-ray crystallography and 1915 Nobel laureate in Physics, came to Copenhagen and excitedly reported that his junior colleague, the Austrian-born chemist Max Perutz, had ingeniously used synthetic polypeptides to confirm the correctness of Pauling's a-helix (see Plate 10). It was a bittersweet triumph for Bragg's Cavendish Laboratory. The year before, they had completely missed the boat in their paper outlining possible helical folds for polypeptide chains.
By then Salvador Luria had tentatively arranged for me to take up a research position at the Cavendish. Located at Cambridge University, this was the most famous laboratory in all of science. Here Ernest Rutherford first described the structure of the atom. Now it was Bragg's own domain, and I was to work as apprentice to the English chemist John Kendrew, who was interested in determining the 3-D structure of the protein myoglobin. Luria advised me to visit the Cavendish as soon as possible. With Kendrew in the States, Max Perutz would check me out. Together, Kendrew and Perutz had earlier established the Medical Research Council (MRC) Unit for the Study of the Structure of Biological Systems.
A month later in Cambridge, Perutz assured me that 1 could quickly master the necessary X-ray diffraction theory and should have no difficulty fitting in with the others in their tiny MRC Unit. To my relief, he was not put off by my biology background. Nor was Lawrence Bragg, who briefly came down from his office to look me over.
I was twenty-three when I arrived back at the MRC Unit in Cambridge in early October. I found myself sharing space in the biochemistry room with a thirty-five-year-old ex-physicist, Francis Crick, who had spent the war working on magnetic mines for the Admiralty. When the war ended, Crick had planned to stay on in military research, but, on reading Schrödinger's What Is Life?, he had moved toward biology. Now he was at the Cavendish to pursue the 3-D structure of proteins for his Ph.D.
Crick was always fascinated by the intricacies of important problems. His endless questions as a child compelled his weary parents to buy him a children's encyclopedia, hoping that it would satisfy his curiosity. But it only made him insecure: he confided to his mother his fear that everything would have been discovered by the time he grew up, leaving him nothing to do. His mother reassured him (correctly, as it happened) that there would still be a thing or two for him to figure out.
A great talker, Crick was invariably the center of attention in any gathering. His booming laugh was forever echoing down the hallways of the Cavendish. As the MRC Unit's resident theoretician, he used to come up with a novel insight at least once a month, and he would explain his latest idea at great length to anyone willing to listen. The morning we met he lit up when he learned that my objective in coming to Cambridge was to learn enough crystallography to have a go at the DNA structure. Soon I was asking Crick's opinion about using Pauling's model-building approach to go directly for the structure. Would we need many more years of diffraction experimentation before modeling would be practicable? To bring us up to speed on the status of DNA structural studies, Crick invited Maurice Wilkins, a friend since the end of the war, up from London for Sunday lunch. Then we could learn what progress Wilkins had made since his talk in Naples (see Plate 11).
Wilkins expressed his belief that DNA's structure was a helix, formed by several chains of linked nucleotides twisted around each other. All that remained to be settled was the number of chains. At the time, Wilkins favored three on the basis of his density measurements of DNA fibers. He was keen to start model-building, but he had run into a roadblock in the form of a new addition to the King's College Biophysics Unit, Rosalind Franklin.
A thirty-one-year-old Cambridge-trained physical chemist, Franklin was an obsessively professional scientist; for her twenty-ninth birthday all she requested was her own subscription to her field's technical journal, Acta Crystallographica. Logical and precise, she was impatient with those who acted otherwise. And she was given to strong opinions, once describing her Ph.D. thesis adviser, Ronald Nor
rish, a future Nobel Laureate, as "stupid, bigoted, deceitful, ill-mannered and tyrannical." Outside the laboratory, she was a determined and gutsy mountaineer, and, coming from the upper echelons of London society, she belonged to a more rarefied social world than most scientists (see Plate 12). At the end of a hard day at the bench, she would occasionally change out of her lab coat into an elegant evening gown and disappear into the night.
Just back from a four-year X-ray crystallographic investigation of graphite in Paris, Franklin had been assigned to the DNA project while Wilkins was away from King's. Unfortunately, the pair soon proved incompatible. Franklin, direct and data-focused, and Wilkins, retiring and speculative, were destined never to collaborate. Shortly before Wilkins accepted our lunch invitation, the two had had a big blowup in which Franklin had insisted that no model-building could commence before she collected much more extensive diffraction data. Now they effectively didn't communicate, and Wilkins would have no chance to learn of her progress until Franklin presented her lab seminar scheduled for the beginning of November. If we wanted to listen, Crick and I were welcome to go as Wilkins's guests.
Dna: The Secret of Life Page 6