|Impact of forty years of advances in chemistry on evolutionary theory|
Presented on September 8, 2003
Roland F. Hirsch
*This presentation reflects the views of the author and does not represent an official position of any unit of the U.S. Department of Energy.
My purpose today is to tell you about the significance of progress in chemistry over the past four decades for the status of Darwinian evolutionary theory. My conclusion—as stated in the abstract —is that advances in chemistry have shifted the focus of the life sciences away from seeking explanations of biological function from origins and toward seeking understanding of function from experiment. Copies of my slides are available, and for those who wish references beyond those in my talk, please send me a note.
Concepts central to evolutionary theory were developed in the first half of the nineteenth century, culminating in the publication of Charles Darwin’s On the Origin of the Species in 1859. The positive reception of this book by the public and the rapid dissemination of Darwin’s ideas reflected a pre-existing receptivity to the ideas, despite the lack of scientific explanations for many of them. The status of Darwinian evolution in science did not grow until the rediscovery of Mendel’s concepts of inheritance in the early 1900s, and the 50th anniversary of Origin passed rather quietly in 1909. Even the public controversy of the 1920s was not about science but about the extra-scientific implications of evolutionary theory as it was then understood.
The Scopes Trial of 1925 exemplifies this. It was about the teaching of a concept of human evolution, not the science of evolution, and I am sure all in this room would have voted to convict Scopes if we had been present on the jury in Dayton, Tennessee. By the way, if you have seen the play or movie Inherit the Wind, forget everything you saw. Every significant historical aspect is portrayed inaccurately.
Progress in genetics and in statistics combined with experimental research led to the codifying of a scientific basis for Darwinian evolution during the 1930s resulting in what is called the Modern Synthesis. From then through the 1950s the influence of evolutionary theory in biology grew rapidly, as scientific evidence accumulated in favor of the Darwinian concepts.
By the time of the centennial of Origin in 1959, Darwinian evolution seemed secure. Noteworthy among the celebrations was one that brought many of the most important biologists of the time to a meeting in the Rockefeller Chapel at the University of Chicago. Led by Julian Huxley they proclaimed the victory of natural selection and its many implications for biological science and beyond.
So here then is the peak of influence of Darwinian evolutionary theory, and the starting point for my analysis of the impact of advances in chemistry. For, as pointed out by Freeman Dyson revolutions in scientific technology lead to discoveries of new things that require new explanations:
And, starting around 1960, several new tools enabled discovery of completely new information about the chemistry of life. While many of these technologies or their analogs were known in 1959, and indeed x-ray diffraction data had led to the model of dna structure published by Watson and Crick in 1953, their application to biology required heroic efforts and very few chemists had access to the necessary instrumentation.
The growth in the importance of chemistry for the life sciences since 1959 must be obvious to all in this room. How many Chemistry Departments are now Departments of Chemistry and Biochemistry! Much of this growth has been enabled by the availability of the new tools for routine use. Where a determination of the structure of a biological macromolecule by x-ray diffraction took years in the 1960s today the complete data set can be obtained in under a day. Where even six years ago we at DOE were proud that our Production Genomics Facility in Walnut Creek California achieved the sequencing of 20 million bases of the human genome at a cost of around $50 million, the facility will sequence more than 2 billion bases in the coming year at the same cost—and they are inviting suggestions of new organisms to sequence.
I want to spend a significant amount of time on the impact of gene sequencing, since it addresses some of the fundamental concepts of evolutionary theory. Here are some of the things that gene sequencing is telling us:
The first item, deletion bias, explains why microbial genomes are so small, given that bacteria and archaea regularly gain large gene segments through interspecies transfers. Yet the size of microbial genomes is very small, many at 1 megabase or less and few above 7 megabases. A study of all the extant complete gene sequences for bacteria available in 2001 came to the conclusion that rather than proceeding by Darwinian random mutation, evolution of these genomes has a substantial bias toward deletion of currently unused gene segments.
It was estimated more than a decade ago that less than five per cent of the human genome codes for genes. The Darwinian explanation was that the non-coding dna is waste from unsuccessful evolutionary experiments. Now we recognize that much, perhaps most of the non-coding parts of the human genome, and the genomes of other multi-celled species, does have function. For one thing, the expression of genes must be regulated, and this requires places for regulatory chemicals (often small peptides) to hook on to the genome as a signal to express a gene. The regulation of gene expression is extraordinarily complex, rather few of the networks are understood at all well. Multiple non-coding segments are needed for regulating most genes. This is especially true for control of the processes involved in development of an organism, before and after birth. 
Scientists have been surprised to learn that the number of genes in a genome that code for proteins does not necessarily determine the complexity of the organism. It was estimated that the human genome would contain at least 100,000 genes. Now it appears that the genome has perhaps 30,000, barely half the number of genes of rice, which has 55,000. Clearly the number of genes does not relate directly to complexity. One of the explanations that chemists have discovered is that, contrary to the expectations of evolutionary theories, many genes code for more than one protein. Factors in the environment determine which is expressed and when.
Let’s now look at the question of vertical inheritance, the essence of Darwinian evolution, versus horizontal transfer of genetic material. The basic concept, as Darwin already stated it in On the Origin of the Species, is of species continually branching away from each other as time increases toward the present. A committee formed by the National Academy of Sciences put it this way as recently as 1998: “The ability to analyze individual biological molecules has added great detail to biologists’ understanding of the tree of life. For example, molecular analyses indicate that all living things fall into three domains—the Bacteria, Archaea, and Eucarya—related by descent from a common ancestor.” Note the clear distinction that is made between each of the three domains of life into which they divide the species.
And what is the reality? In fact genetic material crosses horizontally from species to species across all the domain boundaries. Relationship is not just by descent (in a vertical direction) but also horizontally among species that have no common ancestry but just happen to be in the same place at the same time. In fact, this type of transfer even occurs by a recipient organism “inheriting” genes from a donor organism of a different species in which the donor species was born later than the recipient species. Conceptually it is as if you were to inherit some of your genome from a granddaughter of a unrelated co-worker, born twenty years after you were.
Horizontal gene transfer is an alternative to Darwinian inheritance for explaining anomalous components of a species’ genome. It is detected in a number of ways, one being that a gene in one organism is not present in closely-related species but is present in a distant species, another that the content of bases [C+G versus A+T] is different from the rest of the species’ genome. And it means, contrary to the National Academy’s statement, that the analysis of individual biological molecules does not produce a neat tidy branching from a common ancestor. Ford Doolittle puts it very clearly this way:
Now for an example of why horizontal gene transfer is so important. Photosynthesis occurs in five very different kinds—phyla—of bacteria. How did this come about? Darwinian evolution would suggest that species with similar properties must either be related by descent, or must have gained the property through convergent evolution. Yet based on the chemical data on gene sequences and proteins both explanations are wrong. There are 15 possible trees relating the five phyla. A recent study of representative species in each of the phyla showed that all of the 15 possible trees show some support but none shows even fifteen per cent support. These species gained their photosynthetic ability by horizontal transfer of the key genes, bypassing the Darwinian evolutionary explanations. Understanding photosynthesis by bacteria is essential, for example, to understanding how CO2 is fixed in bodies of water. Yet clearly evolutionary theory is not of much help.
Horizontal gene transfer can be envisioned as occurring through transfer of genetic material by carriers such as viruses, and can be rationalized as plausible for microbial species that are in close contact with each other. So evolutionary theorists thought it would not be found in the more complex multi-celled organisms within the eukaryal domain. Yet many examples of HGT are being found in the Eucaryotic domain:
The second example is especially interesting, as it appears that many parasitic nematodes (small worms) obtain the parasitic function through HGT. Since these worms cause enormous damage to crops, understanding how they become parasitic is of great significance, but is not explained by Darwinian evolutionary theory.
Let’s now go on to some of the impacts of advances in other technologies. Mass spectrometry has in just 15 years become the highest speed technique for studying proteins, thanks to the development of MALDI and ESI, for which half of the 2002 Nobel Prize in chemistry was awarded. Mass spec can measure the expression of large numbers of proteins in a cell under different growth conditions. This kind of analysis, which can be done using a number of techniques in addition to mass spectrometry, is called proteomics. The American Chemical Society last year started a journal devoted to the topic. The reason for the quick growth of proteomics from its conception less than ten years ago is that while knowing the composition of the genome of the organism helps explain function, it is not sufficient, contrary to what was believed forty years ago.
Mass spec also allows studying the ways in which proteins are modified after expression. It turns out that most proteins are modified covalently before they carry out their function. These chemical modifications can now be detected to the point of identifying the sites of specific modifications such as phosphorylation.
The association of proteins into cellular machines is studied in three dimensions using the techniques of structural molecular biology (or biophysics), including small angle x-ray and neutron scattering, x-ray diffraction and cryoelectron microscopy. While these techniques have been known for many decades, it is only since the 1960s that they have become usable for very large molecules and complexes. The x-ray techniques in particular have benefited from the synchrotron light sources at which the first experiments on proteins were done in the early 1970s, experiments that in the last ten years have become rapid and almost routine.
The ribosome is a machine that has some 55 protein molecules as well as several strands of rna in its two subunits in bacteria, and about 75 proteins in eucarya and archaea. It translates the genetic code into protein molecules, a function that is required in all cells. The different proteins in the ribosome are needed to read the code for the amino acid sequence of the protein, find and position each specific amino acid in turn so a peptide bond can be formed extending the protein chain, and initiate the process of folding of the protein.
Other multi-protein complexes are required to carry out most of the essential functions of a cell. Sometimes the proteins each serve to carry out a part of the function of the machine, and sometimes the proteins must be in contact for one of the proteins to assume the proper geometry for its function. Many complexes form in order to move proteins around in the cell, and, before that, to ensure that folding of a protein into its native state occurs correctly. This is especially important because of the extraordinary and unexpected degree to which cells are crowded with proteins and other large molecules.
Chaperone proteins are especially important for these latter functions, and are ubiquitous in all cells. Particularly interesting to chemists are the proteins that control metal transition ions within cells. Some metal ions do not exist at all in cells as the free aquo ion, perhaps not surprisingly, given their strong coordination with sulfur and nitrogen ligands. If zinc or copper were not tightly bound by chaperone proteins that carry them to where they are needed, these metal ions would bind to proteins where they should not bind, disrupting cell function. 
The fact that proteins tend to function as part of multi-protein machines was not intuitive to the Darwinian-trained biologist. Indeed, as cell biologist Bruce Alberts explains:
Thus, forty years ago, at the peak of the Darwinian influence, scientists thought of proteins as diffusing around in a cell and randomly colliding with other molecules to cause the reactions that provide cellular function. This made sense given the one gene -> one protein->one function view of evolutionary biology. The discoveries enabled by the new chemical instrumentation showed that—as Dr Alberts says—proteins in vivo largely are present as parts of machines of ten or more protein molecules. Today much effort is devoted to elucidating the structure and properties of these machines, using the techniques and concepts of chemistry.
The behavior of microbes is increasingly being understood through chemical research. Chemists have helped discover unexpected properties of microbes, properties that are key to understanding microbial function for a multitude of reasons ranging from protection of human health to bioremediation of environmental contamination:
The first discovery startled evolutionary biologists: contrary to the Darwinian notion of competition among species for survival, microbial species cooperate. While a species in pure culture will grow to the limit of the resources available, as Darwinian theory expects, in a natural setting species form a stable community where the numbers of each species stay constant so long as the environment of the community does not change radically.
Chemical signaling is the means that microbes use to maintain a community. The signals could be as simple as the waste chemicals of one species’ metabolism becoming food for a second. In other cases the signals may determine when an activity of a species starts up. The latter communication often falls into the category of quorum sensing, as in the example given in which P. aeruginosa does not cause problems in cystic fibrosis patients until the numbers reach a certain level. 
Surprisingly to evolutionary theorists microbes tend not to reproduce. Generation times in real settings can be days or months where the same species reproduces in hours in pure culture. And, also surprisingly, we have yet to observe a microbe evolving into a different species, despite following as many as a hundred thousand generations in cultures of certain species.
The last item on the above list is about a feature of microbes that is routinely misunderstood, yet one that is of great importance for chemists doing research in drug design. This is that resistance to an antibiotic for treating a pathogen is not due to evolution of a new species of pathogen. When the antibiotic treatment starts, the more susceptible in the population die, leaving behind the less susceptible which require longer treatment. Incomplete treatment courses leave behind these resistant strains. But in many cases the resistant strain actually is less fit than the non-resistant one in the absence of the antibiotic. For example, the normal cells may efficiently transport the antibiotic molecules into the cell, where they are lethal. The resistant molecules may have a deficient transport system so they are not killed by the antibiotic. But they are also less able to transport needed chemicals into the cell, so they die off when the antibiotic is removed, or one targeting a different transport system is used.
Finally, what is the status today of Darwinian evolutionary theory? Here is the assessment of a prominent biologist, editor of one of the leading review journals in biology:
And here is the assessment of a prominent philosopher about why Darwinian evolution continues to be promoted despite its scientific shortcomings:
And here is a cautionary note from Alfred North Whitehead applicable to many theories and certainly to Darwinian evolutionary theory:
Let me sum up with these conclusions:
And thank you for your attention.
This essay is based on a paper presented to the Division of the History of Chemistry at the American Chemical Society National Meeting in New York City, September 8, 2003. The wording has largely been retained from the lecture, except where reference is made to projected slides. The research articles cited are given as examples; the literature is not exhaustively surveyed in this lecture, and many other articles could be included that would be equally useful as examples. Note that the authors of the articles have not been consulted about the interpretation of the impact on Darwinian evolutionary theory presented in this lecture, which in many cases may not have been a consideration in the research projects that they report.
 Darwinian evolutionary theory reached a peak of influence at the centennial of On the Origin of the Species in 1959. Since then advances in experimental technologies have given scientists increasing understanding of the chemistry underlying biological function. Powerful new technologies for DNA sequencing, macromolecular structure determination, microscopy and proteomics, have revealed complexities unanticipated by evolutionary theory. Examples include the recognition of the importance for evolution of horizontal gene transfer (contrasting with the vertical ‘tree of life’ concept), the discovery that proteins usually function as part of complex molecular machines rather than in isolation, and the observation that microbes of different species tend to cooperate rather than compete. It appears that advances in chemistry are shifting interest in the life sciences from seeking historical, evolutionary explanations of life to gaining a chemical understanding of biological function.
 Freeman, Dyson, Imagined Worlds, 1997
 For example: Rapid, accurate, inexpensive gene sequencing; mass spectrometry of proteins; gene microarrays; single-molecule spectroscopy; x-ray diffraction and nuclear magnetic resonance techniques for 3-d structures of proteins; cryo-electron microscopy to characterize protein machines; new means of finding and studying microbes in natural settings
 A. Mira, et al., “Deletional bias and the evolution of bacterial genomes”, Trends in Genetics 17 (2001), 589.
 For example, see: C. Dennis, “Mouse Genome: A Forage in the Junkyard”, Nature 420 (2002), 458.
 E.H. Davidson, et al., “Regulatory gene networks and the properties of the developmental process”, PNAS (USA) 100 (2003), 1475.
 Teaching about evolution and the nature of science. Washington, D.C., National Academy Press, 1998, page 38.
 W. Ford Doolittle, “Phylogenetic classification and the universal tree”, Science 284 (1999), 2124.
 J. Raymond, et al., Science 298 (2002), 1616.
 W.S. Hancock, “The Challenges Ahead”. Journal of Proteome Research 1 (2002), 9.
 An interesting example is found in: M.J. MacCoss, et al., “Shotgun identification of protein modifications from protein complexes and lens tissue” PNAS (USA) 99 (2002), 7900.
 A good
discussion of the structure of the ribosome is found in: A. Yonath, “The
search and its outcome: High-resolution structures of ribosomal
particles from mesophilic, thermophilic, and halophilic
bacteria at various functional states” Annual Reviews of
Biophysics and Biomolecular Structure 31 (2002), 257.
 Bruce Alberts, “The cell as a collection of protein machines: Preparing the next generation of molecular biologists” Cell 92 (1998): 291–294.
 R.M. Donal and J.W. Costerson, “Biofilms: Survival mechanisms of clinically relevant microorganisms” Clinical Microbiology Reviews 15 (2002), 167.
 D.L. Erickson, et al., “Pseudomonas aeruginosa quorum-sensing systems may control virulence factor expression in the lungs of patients with cystic fibrosis” Infection and Immunity 70 (2002), 1783.
 A.S. Wilkins, “Evolutionary processes: a special issue” BioEssays 22 (2000): 1051–1052.
 David Walsh, The Third Millennium: Reflections on Faith and Reason. Georgetown University Press, 1999
 Daly & Cobb,
For the Common Good, 1990, page 36