Oxygen

by Nick Lane

  To place this detail in perspective, here’s a summary of the new evolutionary scheme. LUCA lived in a world scorched by radiation. This in itself suggests that wherever life might have originated, LUCA herself 5 Genes lost more recently are still recognizable, as their sequences have had less time to drift.

  They are called pseudogenes. The completion of the human genome project has given us some idea of how many genes have been lost in this way. A good example is the sense of smell. We once had 900 genes dedicated to detecting smell, but 60 per cent of them are

  ‘broken’ in that no protein can be copied from them. As our primate ancestors took to the trees, vision became more important than smell in the struggle for survival, so many genes for smell could be lost without adverse consequences.

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  must have lived, at least part of the time, in the surface oceans. As the archaea evolved from LUCA (rather than the other way around) the sulphurous, heat-loving extremophiles cannot truly represent the earliest forms of life, as some have argued. On the contrary: if LUCA was metabolically versatile, then she must have lived in a world demanding versatility, and this must have included the surface layer of the oceans.

  The effect of radiation on the ocean surface was far from the barren sterility often portrayed. By splitting water to generate free radicals and hydrogen peroxide, ultraviolet radiation provided an additional source of energy. Hydrogen peroxide was stable enough to build up in shallow seas and lakes before dissociating to produce water and oxygen. Oxygen generated in this way within cells could be captured and stored, using haemoglobin. This oxygen was later released for energy production using cytochrome oxidase. The ubiquitous haem chemical group, a component of cytochrome oxidase and haemoglobin, may even have been the basis for the evolution of the chemically closely related chlorophylls, which could capture light energy and convert it into sugars using variants of the existing respiratory pathways.6

  The first photosynthesizers probably split hydrogen sulphide or iron salts, but as these resources were depleted, in sheltered environments, by oxidative stress (see Chapter 7), selective pressure drove the adaptation to alternatives: hydrogen peroxide and finally water. Once oxygenic photosynthesis had evolved, free oxygen began to build up in the atmosphere and oceans, transforming the world for ever. Yet the ideal oxygen levels for respiratory metabolism remained close to those in which the respiratory enzymes had evolved in the first place. Even today, our own cytochrome oxidase functions best at an oxygen concentration of less than 0.3 per cent of atmospheric oxygen pressure. Our bodies strive to maintain mitochondrial oxygen concentrations at this level.

  While a striking departure in its own right, this new story leads to a conclusion that dovetails with a great deal of recent medical research, in particular the complexities associated with antioxidant treatments. The continuous strain between external and internal oxygen levels lies at the root of many human ailments — a strain that is ultimately worked out in the antioxidant balance of individual cells. The retention of a primordial antioxidant balance is analogous to the salt composition of fluids in the 6 The similarity in the reaction mechanism between cytochrome oxidase and the oxygen-evolving complex has been noted by Curtis Hoganson and his colleagues at Michigan State University, among others; see Further Reading.

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  human body, which still parallels the sea water in which our single-celled ancestors once swam. (J. B. S. Haldane memorably referred to the collection of cells that make up the human body as a sea monster.) The antioxidant balance is too integral to the innermost workings of the cell to behave predictably when subjected to the ruder assaults of medicine. Several of the molecules that we have discussed in this chapter are antioxidants. Catalase breaks down hydrogen peroxide to produce oxygen without generating hydroxyl radicals. Haemoglobins and myoglobins bind oxygen, only releasing it when dissolved oxygen levels have fallen to safe limits. Cytochrome oxidase hoovers up excess oxygen, again without generating free-radical intermediates. Each of these steps regulates intracellular oxygen levels and so attenuates the generation of free radicals. Yet it is quite conceivable that catalase evolved as much to generate oxygen from hydrogen peroxide as to lower toxic levels of oxygen.

  Haemoglobin may well have evolved to hoard oxygen at a time when it was a scarce and valuable resource. Cytochrome oxidase probably evolved as a metabolic enzyme rather than as an antioxidant. Their evolutionary past is built into their modern function, and we ignore it at our peril.

  Such functional opacity highlights the surprising dilemma over the definition of the word ‘antioxidant’, as well as the misguided assumption that molecules evolve for a single purpose. In fact, many so-called antioxidants also serve other purposes, and are tightly woven into the regulatory system of cells. As such, antioxidants maintain oxygen levels within physiological limits, rather than simply mopping up free radicals. This is a crucial distinction. Antioxidant treatments often aim to eliminate free radicals, but by doing so may instead strain the regulatory balance. In thinking about our health, then, it is not enough to consider diseases on their own terms — we must also ask, from an evolutionary perspective, how things got to be that way and what might happen when we fiddle with them. We shall see in the next two chapters just how tightly antioxidants are integrated into the fabric of life. Then, we will examine what this means for our prospects of ameliorating human ageing and disease.

  C H A P T E R N I N E

  Portrait of a Paradox

  Vitamin C and the Many Faces of an Antioxidant

  n apple a day keeps the doctor away says the old adage; but is it true? And if it is true, why? The answer to the first question is Aformulated rather stiffly in our scientific age: a diet containing five 80-gram portions of fruit and vegetables each day reduces the risk of death from heart attacks, stroke and some cancers, especially those of the respiratory and digestive tracts. This is true regardless of our other habits or risks, such as smoking, weight, cholesterol levels and blood pressure.

  Most people currently eat about three portions a day. Several large epidemiological studies have indicated that increasing average consumption to five portions a day might reduce the risk of cancer by 20 per cent and the risk of heart attacks or strokes by about 15 per cent. Health-conscious individuals really do live longer — it is not just that it feels like it, as Clement Freud once remarked. A 17-year study that examined the mortality of 11 000 people recruited through health-food shops, vegetarian societies and magazines, found their mortality rate was half that of the general population (the study was carried out by doctors at the Radcliffe Infirmary in Oxford and reported in the British Medical Journal, not a partisan health-food magazine). Even allowing for the near-intractable methodological difficulties that blight such studies, and my own reluctance to eat fruit, there can be no question that a diet rich in fruit and vegetables is good for you. The problem is rather how to persuade

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  children and adults, especially in northern Europe and the United States, to modify their diets to include five portions a day — a challenge encapsulated by the Europe Against Cancer slogan ‘Take Five!’.

  While the benefits of fruit and vegetables are indisputable, the epidemiology of diet, full as it is of bare associations and correlations, is two-dimensional science. For those of an inquisitive cast of mind, the question why is altogether more interesting and complex. Clearly, fruit and vegetables are filled with goodness; yet, perhaps surprisingly, this is about as much as we know for sure. The depth of our ignorance is conveyed pungently in an article by John Gutteridge and Barry Halliwell:

  “Twenty years of nutrition research have told us that for ‘advanced’ countries the way to a healthy lifestyle is to eat more plants, a concept familiar to Hippocrates. What it has not told us is exactly why.”

  If pressed for an explanation, I imagine that most people would say

  ‘vitamin C’,
‘antioxidants’ and suchlike. The reality is of course far more complicated. The health effects of hundreds, if not thousands, of biologically active compounds isolated from fruit and vegetables have been pored over without real consensus. Given the overwhelming detail, it is not surprising that we tend to fall back on a handful of vitamins that people have at least heard of, but which serve really as surrogates for the consumption of many other compounds. A good example is a study reported by Kay-Tee Khaw and her team at Cambridge in The Lancet in March 2001, which was reported widely and misleadingly by the press: the general projection being that vitamin C lengthens lifespan. In fact, the Cambridge team reported that the risk of death (from any cause) was higher in people with low plasma levels of vitamin C, and conversely, that people with high levels of vitamin C in their plasma were less likely to die within the period studied. The risk of death in people with the highest plasma levels of vitamin C was half that of people with the lowest plasma levels. In the article itself, Khaw and her colleagues were careful to point out that there was no association between vitamin C supplementation and mortality. The association was more generally with dietary intake. And the authors did not discriminate between the amount of vitamin C compared with other factors eaten in the same food at the same time (this is not laziness: asking a specific question often means ignoring superfluous details). No measurements were taken, for example, of plasma levels of vitamin E or beta-carotene. Had they been measured, a similar correlation would almost certainly have been found, for all kinds of antioxidants are abundant in fruit; but that does not mean that they were

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  responsible for the lower number of deaths either. In the Cambridge study, then, plasma vitamin C levels were probably just a surrogate for overall fruit consumption. As to the role of vitamin C itself, we are none the wiser.

  Because it is at once so familiar and so inscrutable, we will use the example of vitamin C in this chapter as a springboard to explore the wider function and behaviour of antioxidants. Although often defined simply as a water-soluble antioxidant, vitamin C illustrates many of the difficulties we face in trying to define an antioxidant. Here is Tom Kirkwood, eminent researcher on ageing at the University of Newcastle, and presenter of the 2001 BBC Reith lectures, giving a vivid depiction of the action of vitamin C:

  When a molecule of vitamin C encounters a free radical, it becomes oxidised and thereby renders the free radical innocuous. The oxidised vitamin C then gets restored to its non-oxidised state by an enzyme called vitamin C reductase. It is like a boxer who goes into the ring, takes a hit to his jaw, goes to his corner to recover, and then does it all over again.

  Kirkwood’s description is not wrong, but it is one-sided. His memorable simplicity conceals a can of worms. The molecular action of vitamin C is as simple and repetitive as flipping a coin, yet the effects are varied, unpredictable and utterly dependent on the milieu in which it operates.

  Just as flipping a coin leads to diametrically opposed outcomes, so too, vitamin C may on the one hand protect against illness and on the other kill tumours, or even people. The food chemist William Porter summed up the conundrum nicely, rising to an anguished eloquence rarely matched in scientific journals: “Of all the paradoxical compounds, vitamin C probably tops the list. It is truly a two-headed Janus, a Dr Jekyll-Mr Hyde, an oxymoron of antioxidants.”

  Few subjects have polarized medical opinion more violently or senselessly than vitamin C. If one man can be held responsible for this division, it is the great chemist, peace advocate and double Nobel laureate Linus Pauling. We will consider Pauling’s life briefly, because his views should not be taken lightly. Neither, we shall see, should they be taken uncritically.

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  It is unfortunate, not least for the memory of Pauling himself, that his legacy should be tainted by his controversial views on vitamin C. No researcher had a more profound impact on the advances of chemistry in the twentieth century. One reviewer of his classic 1939 textbook, The Nature of the Chemical Bond and the Structure of Molecules and Crystals, went so far as to say that chemistry could now be understood rather than being memorized. Pauling was awarded his first Nobel Prize in 1954 for his

  “research on the nature of the chemical bond … and its application to the elucidation of complex substances”. In effect, this meant that he had been awarded the prize for the body of his work over the previous twenty years rather than a specific discovery, a move unprecedented in the history of the Nobel Institute. Yet a continuous thread did run through much of Pauling’s early research — an application of the laws of quantum mechanics to the structure of chemical bonds. Pauling set about calculating the length and the angles of individual bonds, using x-ray diffraction, magnetism and measurements of the heat emitted or absorbed in chemical reactions. From the values he obtained, he went on to plot the three-dimensional structure of complex molecules. One of Pauling’s earliest and greatest contributions to chemistry was the idea of resonance, in which electron delocalization stabilizes the molecular structure (the electron

  ‘spreads out’ in space to dilute the charge density). This feature is critical to the action of vitamin C and other antioxidants, as we shall see.

  By the mid 1930s, Pauling had begun to apply his analytical methods to the structure of proteins. He demonstrated the importance of minute electrical charges (hydrogen bonds) in stabilizing the three-dimensional shape of proteins, and was the first to describe their grand architectural structures, familiar to all students of biochemistry, such as the alpha-helix and the beta-pleated sheet. By the early 1950s, Pauling was turning his attention to the unsolved structure of DNA. In his famous book The Double Helix, James Watson describes the foreboding that he and Francis Crick felt when they heard that the ‘world’s greatest chemist’ was consider-ing the problem of DNA. The pair raced to apply Pauling’s own methods to pip him to the post, and were overjoyed when they realized that, on this occasion, their rival had committed an elementary blunder.

  By now Pauling, spurred on by his indefatigable wife Ava Helen, was becoming increasingly committed to anti-war protests. From 1946, through the 1950s and 1960s, Pauling spoke out about the perils of atomic fallout, in particular the risk of birth defects and cancer. In 1957, he drafted a petition to end nuclear weapons testing, and eventually presented the

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  signatures of 11 000 scientists to the White House, including those of Albert Einstein, Bertrand Russell and Albert Schweitzer. The petition was widely credited with precipitating the Nuclear Test Ban Treaty, in which the United States and the Soviet Union agreed to cease testing nuclear weapons. Pauling was awarded the Nobel Prize for Peace on 10 October 1963, the same day that the Nuclear Test Ban Treaty went into effect.

  Pauling’s anti-war activities inevitably raised the suspicions of the United States government in the early years of the Cold War, when the House Un-American Activities Committee and Senator McCarthy were embroiled in their notorious communist witch-hunt. During the early 1950s, Pauling had been investigated by the FBI and was refused renewal of his passport, receiving the explanation that “Your anti-communist statements haven’t been strong enough”. Only in 1954, when he won the Nobel Prize for Chemistry, and the New York Times brought the controversy to light, was he permitted to travel again. Similar struggles plagued his position at the California Institute of Technology. His funding from the National Institutes of Health (NIH) was cut, along with that of 40

  other scientists, and he was eventually forced to resign from the faculty in 1963. After an interim of several years at the Center for the Study of Democratic Institutions in Santa Barbara, where he devoted himself to the problems of peace and war, he finally took up a chair in chemistry at Stanford University in 1969. There, he pursued his burgeoning interest in

  ‘orthomolecular’ substances, such as vitamin C, which he defined as substances normally
present in the human body and required for life. He went on to establish the Linus Pauling Institute for Orthomolecular Medicine, to which he devoted his remaining years.

  This brief biography must stand as a measure of the man who, in 1970, published the hugely popular book Vitamin C and the Common Cold, in which he claimed that large doses of vitamin C could prevent or cure the common cold. Pauling and his wife practised what they preached, consuming between 10 and 40 grams of vitamin C each day (several hundred times the recommended daily allowance), even adding spoonfuls to their orange juice. Over the next two decades, Pauling’s claims became still more contentious: so-called ‘mega-doses’ of vitamin C could cure schizophrenia and cardiovascular disease, prevent heart attacks and ward off cancer, perhaps adding decades to our life expectancy. Most controversially of all, Pauling and the distinguished Scottish oncologist Ewan Cameron reported that mega-doses of vitamin C, given intravenously, could quadruple the survival time of patients with advanced cancer, even

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  bringing about complete remission in some cases. The medical profession responded to these claims with suspicion, but the Mayo Clinic in Rochester, Minnesota, did at least conduct three small-scale clinical trials to test the effect of vitamin C in advanced cancer. All three trials failed to detect any benefit. Pauling and Cameron argued that the trials had been designed to fail: in particular, that vitamin C had been withdrawn too soon, and was given orally rather than intravenously. In 1989, the NIH

  agreed to review 25 case studies, to be selected by Cameron, for plausible evidence that mega-doses of vitamin C might have important effects in cancer. They concluded, in a letter to Pauling in 1991, that the case studies did not provide convincing evidence of a link.