To the editors:

I learned only recently that when Sir Isaac Newton made his oft quoted comment about having stood on the shoulders of giants, he was not in fact being original.¹ Newton was paraphrasing a reference by the twelfth-century French philosopher Bernard of Chartres to the legend of Orion, the giant-sized hunter in classical mythology who, having been blinded as a punishment, carried the dwarf Cedalion on his shoulders as his guide. Yet regardless of its origins, it is particularly apt that Rosalind Franklin should have invoked this phrase. With a Mars rover named in her honor, a commemorative coin issued by the Royal Mint to mark her hundredth birthday, and an award winning portrayal of her by Nicole Kidman in a hit West End play, Franklin is no longer quite the unsung heroine of DNA that she once was. But there still remains another heroine in this particular story whose role remains very much unsung and upon whose shoulders Franklin herself stood.

In 1938, the physicist Florence Bell took the very first successful X-ray images of DNA, from which she and her supervisor William Astbury proposed an early model for its structure. Although several features of this model were incorrect, Astbury and Bell showed that the bases stack up on top of each other and were able to measure the actual distance between adjacent bases.² This gave James Watson a vital clue when he and Francis Crick began their own work on DNA. But the real legacy of Bell’s work was to show that the regular, ordered structure of DNA could be revealed by the X-ray crystallographic methods that Franklin later put to use.³

In 1941, Bell’s work on DNA ended abruptly when she was summoned for war service. Despite the best efforts of the University of Leeds, she never returned to Astbury’s laboratory but instead married a American serviceman and emigrated to the United States. This came as a blow to Astbury, but in the face of several other challenges, including a lack of funding and facilities in the immediate postwar years, he continued as best he could. In 1951, Astbury’s dogged persistence paid off when his research assistant, Elwyn Beighton, obtained a set of striking new X-ray images of DNA. These showed a cross-shaped pattern that was almost identical to that seen in Photo 51, the image taken a year later by Franklin and Raymond Gosling that is now inscribed on a commemorative fifty pence coin.

In his memoir, The Double Helix, James Watson recalled that when he had first been shown Franklin and Gosling’s photograph, his jaw dropped.⁴ Watson had recognized at once that the striking cross-shaped pattern at the heart of the photograph was characteristic of a molecule with a helical shape. Astbury’s response to Beighton’s earlier images could hardly have been more different. He never published them in a scientific paper, or even presented them at a meeting. Instead, they were filed away and forgotten. As Gareth Williams observes in his book, “A year later and in another place, Beighton’s photographs would have created a sensation. Instead, Astbury did not know what to make of them.”⁵

But what if Astbury had known what to make of Beighton’s image? Might he have gone on to be remembered as the discoverer of the double-helical structure of DNA? This is one possibility considered by Matthew Cobb, author of Life’s Greatest Secret, who has imagined a number of alternative historical routes by which the structure of DNA might have been discovered.⁶

Whilst acknowledging that speculation about counterfactual histories in science is “often typical of late-night bar discussions at conferences,” Cobb maintains that it can nevertheless serve a serious purpose.⁷ Speculation about what might have happened can sometimes be invaluable in challenging assumptions and prejudices about what actually did happen.

The suggestion that Astbury might have solved the double-helical structure of DNA is a good example. Even if Astbury had grasped that DNA had a double-helical shape, this discovery would have offered him no clues about the function of the molecule, in the same way it did for Watson and Crick. They had the crucial insight that a double-helical structure allowed the pairing of complementary bases on opposite strands and that this pairing allowed the molecule to replicate itself and copy genetic information. In a second Nature paper published six weeks later, Watson and Crick proposed what Cobb has called their “brilliant suggestion” about how the molecule might actually carry this information in the first place: “[I]t therefore seems likely that the precise sequence of the bases is the code which carries the genetical information.”⁸

Watson and Crick’s understanding of the structural importance of base-pairing, as well as their “brilliant suggestion” owed much to the work of University of Columbia biochemist Erwin Chargaff. Along with Astbury, Chargaff had been one of the few scientists to sit up and take notice of the discovery by microbiologists Oswald Avery, Maclyn McCarty, and Colin Macleod in 1944 that nucleic acids could confer the property of virulence in bacteria.⁹ Most scientists at this time believed that proteins were the genetic material and had dismissed DNA as being simply a monotonous repeat of the same four bases that did not have sufficient chemical variation to carry genetic information. In the words of the historian Horace Judson, DNA was considered to be merely “the wooden stretcher behind the Rembrandt.”¹⁰

With Avery’s discovery, this began to change. Astbury was so excited by Avery’s work that he described it as “one of the most remarkable discoveries of our time” and he wrote to Avery asking for a sample of DNA so that he could study its structure using X-ray methods.¹¹ Chargaff was equally inspired. Reflecting on his response to Avery’s discovery, he wrote,

I saw before me in dark contours the beginning of a grammar of biology … Avery gave us the first text of a new language, or rather he showed us where to look for it. I resolved to search for this text.¹²

But Chargaff took a different approach than Astbury. His hunch was that this new language was written in the four nucleotide bases that make up DNA. To put this to the test, he wanted to analyze, not its three-dimensional structure as Astbury did, but its chemical composition. By analyzing the quantities of the four nucleotides in DNA obtained from different species, Chargaff made two crucial discoveries. The first was that the ratios of adenine to thymine and cytosine to guanine were often roughly close to one. The second revelation was that the amounts of the four nucleotides showed a significant variation between species. This demolished the idea that DNA was little more than a monotonous repeat of tetranucleotides and suggested that the genetic code might well be carried through the variation in bases.

In his majestic and exhaustive history of molecular biology, The Eighth Day of Creation, Judson has hailed this achievement as being “the discovery that made molecular biology possible and for which we must honor Erwin Chargaff.”¹³ It is not without a certain irony however that this honor fell to Chargaff and not Astbury. For the very technique that had enabled Chargaff to make this discovery had been invented down the road from Astbury’s laboratory at the University of Leeds. And, like Astbury and Bell’s work on DNA, it too had its origins in the study of wool.

To sports fans around the world, the Leeds suburb of Headingley will always be associated with cricket thanks to legendary performances by Sir Ian Botham in 1981 and more recently Alastair Cook. But only a few minutes’ walk from the world-famous test cricket ground, stands a small and ruined stone building. Neither the entourages of fancy dress clad students heading for a night of revelry in the city center nor the passengers who glimpse it from the top deck of the bus are likely to have any idea that a quiet scientific revolution took place here which resulted in a Nobel Prize and the transformation of molecular biology.

Along with an old stone gatepost, this ruin is all that remains of the technical headquarters of the Wool Industries Research Association (WIRA). This was one of several bodies that had been established by the British government at the end of the First World War with the aim of promoting the application of basic science to industrial problems, largely in response to fears that Britain might soon be overtaken by economic rivals such as Germany. Ever since the Middle Ages, when the Cistercian monks at Kirkstall Abbey near the center of Leeds had raised sheep and sold their fleeces to foreign wool merchants, the textile industry had been at the heart of the local economy. Leeds was therefore chosen as the ideal location for WIRA.

In 1938, Archer Martin, a young scientist, arrived from Cambridge and was soon joined by his colleague Richard Synge. At first glance, Martin and Synge’s research may have seemed unlikely to set the scientific world on fire. Their initial aim was to separate and analyze the different amino acids found in wool proteins according to their differential solubility in two solvents. Their first attempts to do this involved using a machine that Martin had built in Cambridge and had brought with him to Leeds. The machine used the countercurrent flow of two different solvents to separate amino acids obtained from acid hydrolysis of wool proteins and was described by Martin as “a fiendish piece of apparatus.” It proved to be an apt description, for its operation was far from a pleasant experience. The operator was required to sit at the machine for a shift lasting four hours while chloroform fumes leaking from the apparatus gradually filled the room. Martin later recalled that when one of the pair came to relieve the other at the end of a shift, he inevitably found himself on the receiving end of a string of expletives from his colleague who was by then suffering the effects of solvent inhalation.¹⁴

The eventual discovery that this machine could be dispensed with therefore came as a great relief. Martin realized that there was no need to have two different solvents flowing past each other. Instead, one could be held immobile on a solid matrix while the other flowed over it. At first this was done using silica gel recovered from packing cases but was then made even easier by using filter paper on which the different types of amino acids separated as distinct purple spots that could be visualized with the staining agent ninhydrin. The method was simple, effective, and, because the separation relied on the distribution, or partition, of amino acids between two different solvents, it became known as partition chromatography.

When it was first presented at a meeting of the Biochemical Society, Martin recalled that partition chromatography using filter paper “raised not a flicker of interest.”¹⁵ Perhaps the audience failed to see how a method of improving the chemical analysis of wool could be of any interest beyond the textile industry. If so, they were to be proven spectacularly wrong. Martin and Synge had done far more than simply invent a new method of separation. They had also conceived of a new idea that would come to define molecular biology. This was that their method might be used not only to determine the composition of amino acids in a protein, but also the precise linear order in which they were arranged—or, as they called it, their sequence.¹⁶

Their early application of this idea to keratin in wool demolished the notion so beloved of Astbury and fellow giants in the field of proteins, Max Bergmann and Carl Niemann, that amino acids in proteins were arranged in neat, regular, repeating mathematical patterns. Building on this development, Martin and Synge now wanted to show that their method could be used to determine the complete sequence of a protein. The problem was that the keratin proteins in wool were simply too large for complete sequencing at this stage. Something smaller was needed and, with the Second World War raging, the ideal candidate had become available.

Gramicidin S was an antibiotic composed of amino acids that was being used by the Soviet Army to treat battlefield casualties; it provided Martin and Synge with the ideal subject on which to test their idea. To help with this work, Synge recruited two young scientists, both of whom would go on to become renowned figures in the field of protein structure. One was the crystallographer Dorothy Crowfoot Hodgkin, who later won the 1964 Nobel Prize in Chemistry for her X-ray studies of vitamin B12 and penicillin, and the other was Frederick Sanger. By that time, Sanger had already started working on identifying the terminal amino acids in insulin. In 1947, when Martin and Synge published the precise order of amino acids in gramicidin S, Sanger became confident that the same feat might be achieved for the entire protein. After the best part of ten years of work, he finally published the complete amino acid sequence of insulin, an achievement which The Times of London likened to Sir Roger Bannister running the first four-minute mile.¹⁷

Sanger’s analysis of insulin hammered the final nail into the coffin of the idea that amino acids were ordered in precise, regular patterns in proteins. But in one of their early papers on partition chromatography, Martin and Synge reflected that “the possible field of usefulness of the new chromatogram is by no means confined to protein chemistry.”¹⁸ Thanks to Chargaff’s work on DNA, this turned out to be a significant understatement.

Although Chargaff had been inspired by Avery’s discovery to turn his attention to DNA analysis, he had faced a formidable obstacle. At the time there was no method available with sufficient precision that could enable him to separate the four nucleotides. But as Williams explains in his book, this all changed when Martin visited Columbia and gave a lecture about partition chromatography.¹⁹

This was Chargaff’s epiphany: Martin and Synge’s method was exactly what he needed for his analysis of DNA.

Why then did it fall to Chargaff, and not Astbury, to take the initiative of applying partition chromatography to the analysis of DNA? Astbury was, after all, working in close collaboration with Martin and Synge and on one occasion had written with delight that “we are rushing together in a mutually helpful manner.”²⁰ But although he declared with excitement “that we are on the verge of something epoch-making in protein studies,” his correspondence at the time betrays a sense that he saw the work at WIRA as being very much subservient to his own X-ray studies of the three-dimensional structure of wool proteins. For Astbury, molecular biology was always rooted in understanding the three-dimensional structure of proteins and DNA.

Equally revealing is an exchange between Astbury and Martin at a symposium held in Leeds in 1946.²¹ Here, Martin showed how his analysis of wool proteins using partition chromatography refuted Astbury’s cherished model that they were composed of a simple pattern of alternating polar and nonpolar amino acids. In summary, Martin offered the blunt conclusion that

the structure of wool is more complicated than has been suggested by Astbury and does not conform to the Bergmann–Niemann hypotheses. Astbury has looked at the structure of proteins “constructively and with the eye of stoichiometric faith.” Lacking this eye and approaching the problem analytically, the present author is unable to find the same degree of order and simplicity.²²

Astbury’s response was to praise Martin as an “outstanding experimentalist,” but this label may not have been entirely complimentary. This formulation suggested that, as a physicist, Astbury may have been asserting an innate intellectual superiority over Martin and Synge. In the television comedy Big Bang Theory, the fictional theoretical physicist Sheldon Cooper similarly regards his roommate Leonard Hofstadter with smug disdain for being merely an experimental scientist. In his response, Astbury went on to regret that he could not

help feeling sad that he [Martin] should confess himself lacking “in the eye of stoichiometric faith.” This is an age of little faith, and it would be a pity were the depression to spread to the proteins just when hopes were beginning to run so high. One thing we can be sure of is that proteins are not a hodgepodge, but are constructed to ordered plans and it is essential to keep this point steadfastly in mind.²³

Yet when Martin and Synge were awarded the 1952 Nobel Prize in Chemistry for the development of partition chromatography, Astbury sent them his warmest congratulations. A few years later in a lecture entitled “In Praise of Wool,” he recalled how, when he had first begun working on wool in the late 1920s, it had been dismissed as being “thoroughly dead, unbelievably dull and unprofitable scientifically, and altogether the kind of protein … that no respectable, aspiring biochemist would touch with a barge pole.”²⁴ How things had changed. For now, he hailed the award of the Nobel Prize to Martin and Synge for their development of partition chromatography as being an “episode to the glory of wool that catches the imagination.”²⁵

By the time Astbury spoke those words, his pioneering work in the application of X-ray crystallographic methods to the study of biological fibers had long established him as a towering figure in protein structure and the emerging science of molecular biology. In an obituary to Astbury, his friend and fellow pioneer of structural biology John Desmond Bernal reflected that “[h]is monument will be found in the whole of molecular biology, a subject which he named and effectively founded.”²⁶ Yet like Orion, the giant in classical mythology, Astbury was also blind. His scientific success had been built on an insistence of the primacy of three-dimensional structure that proved to be mistaken. According to his friend and colleague at Leeds the botanist R. D. Preston, Astbury was more interested in showing the importance of molecular structure to a biological problem than the actual nature of the problem itself. Preston observed that as a result Astbury’s “first flush of enthusiasm soon faded if the answer was not forthcoming in a field which did not bear directly on his own views about protein structure.”²⁷

Astbury’s response to partition chromatography is a good example of Preston’s point. And perhaps Astbury’s greatest oversight was his failure to grasp the enormous potential of partition chromatography when he consigned Beighton’s X-ray images of DNA to a drawer. Astbury’s vision was too limited to recognize that Martin and Synge’s method might be anything more than simply a tool to confirm his own hypothesis about protein structure. And while Orion at least had the dwarf Cedalion sitting on his shoulders to act as his eyes, Astbury had no such guide. This might not have been the case, had Astbury’s pleas to the War Office for Florence Bell to be exempted from war service not fallen on deaf ears. With Bell’s formidable intellect to challenge the limitations of his vision, he might have been alerted not only to the importance of Beighton’s photographs for revealing the structure of DNA, but also to the possible application of Martin and Synge’s method to unravel its chemical structure. Then it might well have been Astbury, and not Chargaff who, in Judson’s words, made “the discovery that made molecular biology possible.”²⁸