Pauling, Zuckerkandl and the Molecular Clock

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Dr. Emile Zuckerkandl, 1986.

In 1963, a year after first publishing their ideas on the study of molecules as indicators of evolutionary patterns, Emile Zuckerkandl and Linus Pauling continued to explore what they felt to be a very promising thread of inquiry.

Specifically, the two joined in arguing that the molecular clock method – as they had since termed it – might be used to derive phylogenies (or evolutionary trees) from essentially any form of molecular information. This position was further explicated in “Molecules as Documents of Evolutionary History,” an article published in Problems of Evolutionary and Industrial Biochemistry, a volume compiled in 1964 on the occasion of Soviet biochemist Alexander Oparin’s 70th birthday.

Zuckerkandl and Pauling’s most influential work on this subject was first put forth that same year, in a paper that they presented together at the symposium “Evolving Genes and Proteins,” held at the Rutgers University Institute of Microbiology. The talk, formally published a year later and titled, “Evolutionary Divergence and Convergence at the Level of Informational Macromolecules,” classified molecules that occur in living matter into three groups. Each of these groups was identified according to new terms that the pair had developed that were based on the degree to which specific information contained in an organism was reflected in different molecules. These three categories were:

1.Semantophoretic Molecules (or Semantides), which carry genetic information or a transcript of it. DNA, for example, was considered to be composed of primary semantides.

2. Episemantic Molecules, which are synthesized under control of tertiary semantides. All molecules built by enzymes were considered episemantic.

3. Asemantic Molecules, which are not produced by the organism and do not express (directly or indirectly) any of the information that the organism contains. In their discussion, Zuckerkandl and Pauling were quick to point out that certain asemantic molecules may shift form. Viruses, for example, can change form when integrated into the genome of the host; so too can vitamins when used and modified anabolically.

Semantides were considered most relevant to evolutionary history, but the term never caught on in biology, paleontology, or other allied fields relevant to the study of evolution. Nonetheless, whatever the nomenclature, the “semantides” that Zuckerkandl and Pauling wanted to investigate – DNA, RNA, and polypeptides – proved indeed to be precisely the treasure trove of information on evolutionary history that the duo had hoped would be the case.


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A figure from the French translation of Zuckerkandl and Pauling’s 1964 paper.

Fundamentally, Zuckerkandl and Pauling aimed to elucidate how one might gain information about the evolutionary history of organisms through comparison of homologous polypeptide chains. In examining these substances, the researchers sought specifically to uncover the approximate point in time at which the last common ancestor between two species disappeared. In essence, it is this approach that we speak of when we use the terms “molecular clock” or “evolutionary clock.”

Zuckerkandl and Pauling argued that, by assessing the overall differences between homologous polypeptide chains and comparing individual amino acid residues at homologous molecular sites, biologists and paleontologists would be better equipped to evaluate the minimum number of mutational events that separated two chains.

With this information in hand, researchers would thus be empowered to exhume the details of evolutionary history between species, as inscribed in the base sequences of nucleic acids. This set of data, they believed, would hold even more useful information than would corresponding polypeptide chain amino acid sequences, since not all substitutions in the nucleotides would be expressed by differences in amino acid sequence.


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A table from Zuckerkandl and Pauling’s 1965 Bruges paper.

As their work moved forward, Pauling and Zuckerkandl published another paper, 1965’s “Evolutionary Divergence and Convergence in Proteins.” This publication appeared in Evolving Genes and Proteins, a volume that emerged from a conference that the two had attended in Bruges, Belgium.

By this point, the duo’s idea of the molecular clock, or “chemical paleogenetics,” had elicited opposition from organismal evolutionists and taxonomists, as well as some biochemists. Now referring to their “semantides” simply as informational macromolecules, Zuckerkandl and Pauling used the 1965 meeting to argue strongly against their skeptics. Zuckerkandl chided

Certainly we cannot subscribe to the statement made at this meeting by a renowned biochemist that comparative structural studies of polypeptides can teach us nothing about evolution that we don’t already know.

Pauling likewise added that

Taxonomy tends, ideally, not toward just any type of convenient classification of living forms (in spite of a statement to the contrary made at this meeting).

Directly challenging those present who were attempting to discredit the idea of the molecular clock, the pair insisted that taxonomy tended toward a phyletic classification based on evolutionary history. Since the comparison of the structure of homologous informational macromolecules allowed for the establishment of phylogenetic relationships, the Zuckerkandl-Pauling studies of chemical paleogenetics therefore had earned a place within the study of taxonomy. This, they argued, was true both as a method of reinforcing existing phyletic classifications and also of increasing their accuracy. Specifically, the two claimed that

The evaluation of the amount of differences between two organisms as derived from sequences in structural genes or in their polypeptide translation is likely to lead to quantities different from those obtained on the basis of observations made at any other higher level of biological integration. On the one hand, some differences in the structural genes will not be reflected elsewhere in the organism, and on the other hand some difference noted by the organismal biologist may not be reflected in structural genes.

Indeed, it was these early observations, coupled with additional work conducted by those scientists who took their ideas seriously, that allowed for the development of a successful measure of rates of evolutionary change over time. Without these data, modern paleontologists, physical anthropologists, and geneticists would not be able to accurately determine evolutionary histories. Today, this technique has been systematized and specialized in the field of bioinformatics, which is now foundational to many studies in both biology and medicine.

The taxonomic purpose of the molecular clock, however, was only a byproduct of Zuckerkandl and Pauling’s main ambitions in studying paleogenetics: to better understand the modes of macromolecular transformations retained by evolution; to elucidate the types of changes discernible in information content; and – most importantly for Pauling – to identify the consequences of these changes for a given organism.


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Linus Pauling, 1992.

Despite its considerable potential, the work of Zuckerkandl and Pauling, though conducted at such a critical juncture in a nascent field, was largely forgotten as recently as the early 1990s. In fact, in Allan Wilson and Rebecca Cann’s 1992 article, “The Recent African Genesis of Humans,” it was implied that the concept of the molecular or evolutionary clock was first developed and employed by a Berkeley anthropologist, Vincent Sarich. Sarich had collaborated with Wilson in 1967 to estimate the divergence between humans and apes as occurring between four to five millions years ago.

Pauling was still alive in 1992, and seeing this article he duly wrote to the editor of its publisher, Scientific American, pointing out that that, in fact, he and Zuckerkandl had, in 1962, issued their own estimate of the disappearance of the last common ancestor of gorilla and man. Zuckerkandl and Pauling’s calculations had yielded a divergence at about 7.6 million years before present, which Pauling pointed out was much closer than Sarich’s figure to the more recent estimates of divergence determined by Sibley and Ahlquist in 1984 and 1987. Notably, Pauling and Zuckerkandl’s estimate continues to remain closer to more contemporary notions of 8 to 10 million years.

Today, Emile Zuckerkandl and Linus Pauling are remembered as having first championed the notion of the molecular clock, even if many of the details now deemed as fundamental still needed to be ironed out by an array of scientists who followed. Regardless, as in so many other areas of science, Pauling proved once more to be on the ground floor of a new discipline. This was an academic venture that continued also to serve the younger Zuckerkandl well, as he continued on through a prolific career in science that concluded with his passing in 2013.

Molecules as Documents of Evolutionary History

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Linus Pauling and Emile Zuckerkandl, 1986.

[Part 1 of 2]

“Of all natural systems, living matter is the one which, in the face of great transformations, preserves inscribed in its organization the largest amount of its own past history. We may ask the question: Where in the now living systems has the greatest amount of their past history survived, and how it can be extracted?”

-Linus Pauling and Emile Zuckerkandl, 1964

Austria-born Emile Zuckerkandl fled to Paris with his parents to escape the Nazi occupation of his homeland when he was only sixteen. Twenty years Linus Pauling’s junior, this extraordinary scientific mind was nurtured through the study of the biological sciences at the University of Illinois in the United States, and the Paris-Sorbonne University in France. During the course of these studies, Zuckerkandl developed a particularly keen interest in the molecular aspects of physiology and biology, and is now regarded to have been a major contributor to both fields.

Zuckerkandl first met Linus Pauling in 1957, a time period during Pauling himself was spearheading the new study of molecular disease, following his earlier, groundbreaking discovery of the genetic basis of sickle cell anemia. Two years later, Zuckerkandl joined Pauling as a post-doc in the Chemistry and Chemical Engineering Division at the California Institute of Technology. Once arrived, Zuckerkandl was encouraged by Pauling to study the evolution of hemoglobin, a research project that was informed by a basic and crucial assumption that the rate of mutational change in the genome is effectively constant over time.

This assumption was something of a hard sell for biologists at the time, due to the prevailing belief within the discipline that mutations were effectively random. Undaunted, Pauling and Zuckerkandl moved forward with their project and ultimately created a model that, over time, ushered in major changes to the conventional wisdom.


The Pauling-Zuckerkandl project was first revealed to the world in a co-authored paper that appeared in 1962’s Horizons in Biochemistry, a volume of works written and dedicated to the Nobel winning physiologist Albert Szent-Györgyi. The paper, titled “Molecular Disease, Evolution, and Genic Heterogeneity,” is regarded today as a foundational work in its application of molecular and genetic techniques to the study of evolution.

At the time that Pauling and Zuckerkandl wrote this first joint paper, Pauling was, as the title of the work might suggest, very interested in molecular disease; so much so that he was thinking about molecular disease and evolution as occupying two sides of the same coin. Defining life as “a relationship between molecules, not a property of any one molecule,” Pauling believed that disease, insofar as it was molecular in basis, could be defined in exactly the same way. Since both evolution and molecular disease were merely expressions of relationships between molecules, the distinction between the two became blurred for Pauling, who felt that the mechanism of molecular disease represented one element of the mechanism of evolution.

In their paper, Pauling and Zuckerkandl outlined their point of view as follows:

Subjectively, to evolve must most often have amounted to suffering from a disease. And these diseases were of course molecular. …[T]he notion of molecular disease relates exclusively to the inheritance of altered protein and nucleic acid molecules. An abnormal protein causing molecular disease has abnormal enzymatic or other physicochemical properties. Changes in such properties are necessarily linked to changes in structure. The study of molecular diseases leads back to the study of mutations, most of which are known to be detrimental. A bacterium that loses by mutation the ability to synthesize a given enzyme has a molecular disease. The first heterotrophic organisms suffered from a molecular disease, of which they cured themselves by feeding on their fellow creatures. At the limit, life itself is a molecular disease, which it overcomes temporarily by depending on its environment.

These assertions – in particular that “life itself” was a molecular disease – were so strange and seemingly outrageous that many biologists dismissed the paper outright. However, the basic idea that the pair was considering simultaneously sparked the interest of many other scientists who willing to entertain the consequences of this mode of thinking. If Pauling and Zuckerkandl were right, then every vitamin required by human beings stood as a witness bearing testimony to the molecular diseases that our ancestors contracted hundreds of millions of years ago. These “diseases” would manifest negative symptoms only when the curative properties of the nutrients gained from our natural environment through food and drink proved insufficient to dampen their effects.


This line of reasoning provided the impetus for Pauling and Zuckerkandl to begin examining differences between the genetic code as a tool to better understand evolutionary history. Though they did not call it “the molecular clock” at the time, the concept served as the foundation for the genetic analysis of species over long periods of evolutionary history.

Fundamental to Pauling and Zuckerkandl’s argument was the notion that there was no reason to place molecules at specific points “higher” or “lower” on an evolutionary scale. Horse hemoglobin, for example, is not less organized or complex than is human hemoglobin, it is simply different. The same is true for the genetic information contained in the hemoglobin of humans as compared to other primates, such as gorillas.

As Zuckerkandl examined differences of these sorts, he was not surprised to find that the peptide sequences of a gorilla were more similar to a human’s than a horse’s were to either. He and Pauling then began to consider how these sequences must be indicative of the millions of years of separate evolution that had passed since the disappearance of the last common ancestor linking horses and primates.

While compelling in its own right, for Pauling the chief purpose of understanding these differences was to discern crucial information about the human condition and to define the parameters of optimal health. Indeed, Pauling fundamentally believed that an improved understanding of the transitions that the genetic code had undergone would ultimately reveal new and effective treatments for molecular diseases.


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Table I from Pauling and Zukerkandl’s 1962 paper.

To demonstrate the efficacy of their methodology, Pauling and Zuckerkandl calculated the number of genetic differences that exist between the alpha and beta chains of hemoglobin peptide sequences in horses, humans, and gorillas, which they then used to determine the time that had elapsed since the erasure of the last common ancestor linking these species. In seeming support of their claims, the authors found that their figures matched pretty closely with data uncovered by paleontologists.

Despite this, the impact of Pauling and Zuckerkandl’s paper dissipated pretty quickly. For other established figures in the field, Pauling and Zukerkandl had failed to prove the essential assumption that evolution should proceed with relative uniformity over time. Lacking a clear reason to accept that genetic change occurs at a constant rate, there was no compelling reason to believe that Pauling and Zuckerkandl’s molecular clock should give an accurate picture of evolutionary history.

Nonetheless, the idea of the molecular clock found a degree of traction among biologists who valued its potential to corroborate and increase the accuracy of existing phylogenetic assignments. And as we’ll discuss in our next post, Pauling and Zuckerkandl continued to explore their ideas, eventually building a body of work that came into far greater favor a few decades later.

Emile Zuckerkandl, 1922-2013

Pauling and Zuckerkandl in Japan, 1955.

Pauling and Zuckerkandl in Japan, 1986.

We close out our posting schedule for the year on a melancholy note with this remembrance of the life of Emile Zuckerkandl, who passed away on November 9th at the age of 91.

Zuckerkandl was born in Vienna, Austria on July 4, 1922. His family was of Jewish descent and active in the scientific, artistic, and political culture of the time. His father, Frederick, was a biochemist and his mother, Gertrude, was a portrait and landscape artist whose father, Wilhelm Stekel, was an anatomist. His paternal grandfather, Emil Zuckerkandl, was also an anatomist – one of great prominence – whose wife, Berta, was a journalist and art critic. Berta also hosted salons in Vienna attended by all manner of literary, musical and artistic figures.  She likewise used her position to speak out against Nazi occupation and militarization, a stance that forced the Zuckerkandls to flee from Austria into France in 1938 and to later to Algiers.

The family’s flight from Vienna briefly disrupted educational pursuits for Emile, who was attending high school at the time. He finished school in Paris before going to university in Algiers where he studied medicine. While there – as he would relate in a 1996 interview with Gregory Morgan at the Dibner Institute – “I came to understand that it was biology that I was interested in, more than medicine.” He was soon expelled however, a victim of the Vichy’s anti-Jewish laws that extended to colonial Algiers. By this point, the only schooling still available to him was at the music conservatory, where he studied piano.

Emile and Berta Zuckerkandl, ca. 1930. Image courtesy of the Austrian National Library.

Emile and Berta Zuckerkandl, ca. 1930. Image courtesy of the Austrian National Library.

While the family had escaped the dangers of residence in Austria and France, Emile’s outspoken grandmother was still not completely safe and so she sought to go to the United States. During this time, Albert Einstein took an interest in the young Zuckerkandl and helped him to obtain a scholarship to study in the U.S. The Zuckerkandls ultimately did not need to come to the United States as the Nazis were soon defeated, thus rendering France safe for the family once again. Nonetheless, Emile spent one year at the Sorbonne in Paris before taking advantage of the scholarship that Einstein had helped him to obtain.

Once stateside, Zuckerkandl attended the University of Illinois where, in 1947, he obtained a master’s degree in physiology, spending summers researching at the Marine Laboratory at Woods Hole in Massachusetts. He returned to the Sorbonne afterwards and earned his PhD in biology. Following that, he spent ten years at the Roscoff Biological Station in western France – the country’s largest marine laboratory – where he carried out biochemical research.


Emile and Jane Zuckerkandl, October 1970.  Image courtesy of the Esther M. Lederberg Collection.

Emile and Jane Zuckerkandl, October 1970. Image courtesy of the Esther M. Lederberg Collection.

In 1958, while in Paris, Zuckerkandl arranged to meet with Linus Pauling. The two had been introduced through Alfred Stern, an Austria-born professor of philosophy at Caltech and a friend of the Zuckerkandl family. The following year Pauling recommended Zuckerkandl for a post-doc position at Caltech, stressing his relationship with Stern as well as with Charles Metz, who had received his PhD in biology from Caltech and was the brother of Emile’s wife, Jane.

Buoyed by Pauling’s initial recommendation and continued support, Zuckerkandl spent the next five years at Caltech. While there, Zuckerkandl propelled important work on what would become known as the molecular clock. The project was spurred by a suggestion that Pauling made to Zuckerkandl; that he compare the hemoglobin protein sequences of humans and gorillas to trace their evolutionary development. As Zuckerkandl told Morgan, “When Dr. Pauling made this suggestion to me, I was not too happy,” as he wished to continue his own line of research. But “later I understood how right Dr. Pauling was… At first I did not know how lucky I was!” Indeed, the molecular clock and molecular evolution would form the foundation of much of Zuckerkandl’s scientific career.

In 1961 Zuckerkandl, on Pauling’s recommendation, began working in Walter A. Schroeder’s lab, one of the few in the world researching protein sequences at that time. Zuckerkandl proposed that Schroeder coauthor the publication of his evolution work, but Schroeder refused due to his own creationist views. Pauling came to the rescue by suggesting that he coauthor with Zuckerkandl instead, but stipulated that they “should say something outrageous” since the piece was slated to appear in a prominent collection honoring the birth of Albert Szent-Györgyi.

The result was a classic 1962 paper, “Molecular Disease, Evolution and Genetic Heterogeneity,” in which Zuckerkandl and Pauling first postulated the idea of the molecular clock. As Zuckerkandl told Giacomo Bernardi in 2012, Pauling was more focused on his peace work at the time and his involvement with the paper was relatively tangential. Burdened by a crush of other obligations, Zuckerkandl’s coauthor agreed that he would “make some changes that would be moderate, and then the paper would come out as I conceived it.”

Morgan described the basics of the paper and its impact in a 2001 essay.

The molecular clock hypothesis, as it came to be known, proposed that the rate of evolution in a given protein molecule is approximately constant over time. More specifically, it proposed that the time elapsed since the last common ancestor of two proteins would be roughly proportional to the number of amino acid differences between their sequences. The molecular clock, therefore, would not be a metronomic clock – that is, its ‘ticks and tocks’ would not be uniform – but would instead be a clock based upon random mutation events. In practice, a molecular clock would allow biologists to date the branching points of evolutionary trees.

The molecular clock hypothesis, while rarely cited among Pauling’s most important discoveries, has proven to be very influential. The UC Berkeley biologist Alan Wilson claimed that the molecular clock is the most significant result of research in molecular evolution. In his book Patterns of Evolution, Roger Lewin describes the molecular clock as ‘one of the simplest and most powerful concepts in the field of evolution.’ Francis Crick, co-discoverer of the structure of DNA, called the molecular clock a very important idea that turned out to be much truer than most thought when it was proposed.


For the remainder of his time in Pasadena, Zuckerkandl continued to develop his ideas, and in this was assisted in the laboratory by his wife, Jane. Yet he was also eager to return to France, fueled in part by continued uncertainty surrounding his funding at Caltech. Pauling tried to help by recommending Zuckerkandl for a position in the anthropology department at the University of California – San Diego and by bringing back to Zuckerkandl contact information for scientists in Europe. Upon returning to Europe for a conference in 1962, Zuckerkandl wrote to Pauling, “It was as exciting, after three years of life in California, to rediscover Europe as it had been to first discover America.” Soon afterwards, he obtained a position at a new research facility in Montpellier, France – the only hitch was that he needed to wait three years for it to be built. In the meantime, Zuckerkandl expressed worry to Pauling that his work on gorilla hemoglobin would soon “be out of date” because of the interruption posed by his return to France.

Once Zuckerkandl and his research materials had finally arrived in Montpellier in 1965, new problems arose. He told Pauling, “I did not foresee the particular kind of obstacles that I find in my way. Bureaucracy is taking over the direction of our laboratory.” (In a 1971 letter, when asking how Pauling had been such a “great fighter” for peace and science, Zuckerkandl concluded that it could only have been because Pauling had “not tried to found a research department in France.”)  This manner of struggle proved a defining feature of Zuckerkandl’s years at Montpellier and it was not long before he and Pauling began seeking out avenues to reunite, both through short visits and Zuckerkandl’s standing invitation for Pauling to come to Montpellier as a visiting researcher. A bright spot occurred in 1970 as Zuckerkandl was offered the editorship of the new Journal of Molecular Evolution by Conrad Springer.

Zuckerkandl with Ewan Cameron, New Years Day, 1979. Image courtesy of the Esther M. Lederberg Collection.

Zuckerkandl with Ewan Cameron, New Years Day, 1979. Image courtesy of the Esther M. Lederberg Collection.

Zuckerkandl eventually decided to leave France and return to the United States. In 1975 Pauling offered Zuckerkandl lab space at the Linus Pauling Institute of Science and Medicine, but by then he was already at Woods Hole where he hoped details might get resolved at his old position and he could return to Montpellier. A bit later, when Zuckerkandl decided to take Pauling up on his offer, Pauling had to turn him down due to the Institute’s mounting financial problems. Zuckerkandl stayed at Woods Hole and had assumed a visiting professorship at the University of Delaware when, in 1976, Pauling offered him a year-long non-resident fellowship.


LPISM Newsletter, 1977.

LPISM Newsletter, 1977.

Zuckerkandl quickly moved up the ranks at the Institute. At the end of his fellowship, Pauling recommended that he be sponsored for a two year visa and become Vice Director of the organization. By 1979 Zuckerkandl held the additional positions of Vice President and Research Professor, assuming responsibility for carrying out his own work on vitamin C and cancer while also helping Pauling with administrative duties. By the end of the year, Zuckerkandl was taking on even greater administrative responsibilities from Pauling, including budgeting, fund raising, and finding a new home for the Institute.

During this period, Zuckerkandl’s management skills came to the fore. He immediately set out to establish priorities for research, setting a “cutting off point” in time after which any new projects would be “provisionally abandoned.” He also dealt with the storm of controversy caused by the firing of Institute President Art Robinson and Robinson’s statement in Barron’s that high doses of ascorbate led to tumor development in mice. (In a memo to Pauling, Zuckerkandl noted that “the data do not appear to authorize Art’s statement’s” which were most likely based on a “scatter of the experimental points” and nothing more definite.)

The following year, Zuckerkandl was made President and Director of the Institute. Funding continued to be a problem, with each advance seemingly countered by a setback. Typically, at about the same time that the Institute agreed to pay a settlement to Robinson, Zuckerkandl also oversaw the opening of the Armand Hammer Cancer Research Center, with support from its namesake. In 1985, to show his dedication to the Institute, Zuckerkandl deferred a raise that he had been offered until it could be afforded.

However, in 1992 Zuckerkandl’s contract was not renewed. By then the Institute was in a dire financial position and needed to make some very difficult decisions concerning staffing and programs. Nonetheless, Zuckerkandl remained in the building through a lease agreement between LPISM and his new Institute of Molecular Medicine. He also invited many LPISM staff, some of whom had also been laid off, to join him in his fledgling enterprise.


Emile Zuckerkandl, 1993.

Emile Zuckerkandl, 1993.

When not managing a research institute, Zuckerkandl continued to produce work on molecular evolution. In 1997 and 2007, continuing a career-long debate over the relationship between molecular evolution and natural selection, Zuckerkandl addressed molecular diseases and “junk DNA” to argue against the neutral theory of molecular evolution. Beginning in the late 1960s, Motoo Kimura, Jack King, Thomas Jukes, and others had used the molecular clock idea to suggest that evolutionary change occurs not because of natural selection, but due to neutral random mutations and allele drift. Zuckerkandl disagreed, arguing that natural selection and the molecular clock were compatible.

Zuckerkandl not only argued against what he saw as misapplications of his own ideas, but also against broader cultural attacks on science. Beginning in the late 1980s, he began to speak out against creationists, telling Pauling in a 1991 memo that “since in this country they seriously threaten education and culture, a response to them needs to be made.”  As Zuckerkandl saw it, the threat was not only cultural but also involved the health of humanity and of the Earth. In a 1991 draft titled “Genetic and Esthetic Winter,” which he shared with Pauling, Zuckerkandl proposed the need for more control of population growth to curb human impact on the environment, an imperative that he saw as being impeded partly by religious beliefs.

Zuckerkandl also turned his gaze toward enemies within the academy, attacking the application of social constructivism to science. In a 2000 paper, he argued that while scientific discoveries may be socially determined, the “mature” results that lead to practical applications, especially in technology and medicine, is “evidence for independence” of science from similar influences.

But Zuckerkandl did not maintain a strict scientism. In “Genetic and Esthetic Winter,” he described the resilience of science, but also its limitations. When speaking about threats to environmental stability, he stated,

In this situation, curiously, scientific culture is perhaps the least threatened province of culture. Yet knowing and knowhow are not enough. Perceiving and feeling cannot be replaced by engineering. We can’t be satisfied with preparing a future in which the environment will be enriching only for the vocation of engineers.

Zuckerkandl’s health steadily deteriorated as he advanced in age. Over time, a troublesome back proved to be especially problematic. He died on November 9, 2013 in Palo Alto, California, the victim of a brain tumor.

The Theory of the Molecular Evolutionary Clock

Dr. Emile Zuckerkandl, 1986.

Dr. Emile Zuckerkandl, 1986.

It thus appears possible that there would be no evolution without molecular disease.”
-Linus Pauling. “Molecular Disease, Evolution and Genic Heterogeneity,” 1962.

In the early 1960s, Linus Pauling and Emile Zuckerkandl, a French postdoctoral fellow who had arrived at Caltech in 1959, began researching the characteristics of hemoglobin extracted from a number of different species of animals. Zuckerkandl used a technique called fingerprinting, a process taught to him by a Caltech graduate student named Richard T. Jones, to create patterns of the amino acid sequences in each hemoglobin molecule.

[In her Master of Science thesis (pdf link), Dr. Melinda Gormley described fingerprinting, which was invented by the English chemist Vernon Ingram, as “a two-step process, [that] utilizes paper electrophoresis and paper chromatography. It produces splotches on paper at various locations; each mark corresponds to a peptide (two or more linked amino acids).”]

Once patterns had been prepared for several species, Pauling and Zuckerkandl compared them two at a time, and it was from the results of these comparisons that the theory of the Molecular Evolutionary Clock was developed.

The Molecular Clock differs from other evolutionary theories in that it tracks the evolution of a molecule rather than the evolution of a species. The theory states that, every so often, a mutation occurs in a given hemoglobin molecule. Generally speaking, this mutation is the source of a molecular disease, but will not cause any significant change to any organism other than its host.

Occasionally, however, a mutation will cause a lasting alteration to the molecule, and as the organism with the altered molecule reproduces, the change becomes permanent. More alterations of this nature can then occur on top of the original modification, thus resulting in even more differences in those hemoglobin molecules that have descended from the mutated original.

It is these differences that Pauling and Zuckerkandl were interested in when they compared the fingerprint patterns of different species, and their research led to an important breakthrough. As the duo compared more and more fingerprint patterns in a wider range of combinations, they observed that the number of differences between fingerprints lessened as the two species became more closely related. Pauling later stated that

[Zuckerkandl] found that in the beta chain of the human and the beta chain of the horse, for example, 20 of the 146 amino acids are different; but with human and gorilla, only one is different. It is the same amount of difference, just one amino acid residue, as between ordinary humans and sickle cell anemia patients, who manufacture sickle-cell-anemia hemoglobin.

From there Pauling and Zuckerkandl proposed that the comparative-fingerprinting method could be used to speculate as to how long ago any two species deviated from a common ancestor. Even more specifically, they reached the conclusion that one amino acid would be substituted every eleven to eighteen million years for any given species.

The evolutionary theory of the Molecular Clock was not readily accepted by scientists because it proposed a constant rate of evolution. However, it’s importance has now been noted and more research has been done on Pauling and Zuckerkandl’s original work. See, for instance, the very thorough examination conducted by Dr. Gregory J. Morgan in his 1998 paper “Emile Zuckerkandl, Linus Pauling, and the Molecular Evolutionary Clock, 1959-1965.” [pdf link], as well as Naoyuki Takahata’s “Molecular Clock: An Anti-neo-Darwinian Legacy,” [Genetics, (May 2007) 176: 1-6; not freely available online] which concludes that “a molecular clock is a most remarkable manifestation and a tribute from nature to anyone who studies evolutionary biology.”

For more information on Pauling’s hemoglobin work, please visit the website It’s in the Blood!  A Documentary History of Linus Pauling, Hemoglobin and Sickle Cell Anemia, and for more on Linus Pauling, check out the Pauling Online portal.