Oxygen
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This means that we need to know which fragments are diagnostic of hydroxyl radical attack, and what proportion of the overall molecular wreckage they constitute.
One modified DNA building block that probably does reflect hydroxyl radical attack is 8-hydroxydeoxyguanosine (8-OHdG), a chemically modified form of deoxyguanosine, the G in the four-letter DNA code. Ames and his team measured the concentration of this molecule in the urine of rats, and then extrapolated from the results to calculate the likely number of hydroxyl radical ‘hits’ to DNA in each cell of the rat’s body. They concluded that there are as many as 10 000 ‘hits’ to the DNA of each cell every day, although most of these are presumably repaired immediately
— hence the 8-hydroxyguanosine excreted in the urine. More recent studies have examined the equivalent rate in people. The rate seems to be lower than in rats, but may still approach several thousand hits per cell each day. This is several orders of magnitude lower than the projected 4 million hydroxyl radiacals estimated to be produced in a cell each day, but bear in mind that the figure of 10 000 represents hits to DNA alone, and does not include potentially damaging hits to lipids in cell membranes or to proteins, which make up a far greater part of the cell than does DNA.
Despite the wide margins for error, the fact that hydroxyl radicals are produced from both breathing and radiation allows a direct comparison to be made, at least in principle. James Lovelock used figures similar to
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those cited above to calculate the equivalent dose of radiation that we absorb by breathing. He estimates that the damage done by breathing for one year is equivalent to a whole-body radiation dose of 1 sievert (or 1 joule energy per kilogram). As a typical chest x-ray delivers 50 micro-sieverts of radiation, breathing for a year would seem to be 10 000 times more dangerous than a chest x-ray, or 50 times as dangerous as all the radiation that we normally receive from all sources in the course of an entire lifetime.
These figures, while impressive, are somewhat disingenuous. For a start, we do not know whether these ‘hits’ to DNA are to working genes, or to inactive genes, or to ‘junk’ DNA, which does not code for anything and, in fact, comprises the bulk of human DNA. In addition, there is one major difference between radiation and breathing — the starting point. Radiation produces reactive hydroxyl radicals from water immediately, and they have a random distribution throughout the cell. Because our normal exposure to radiation is low we have not evolved to deal with this pattern of distribution or immediate reactivity. Respiration, on the other hand, produces mainly superoxide radicals, which are altogether less reactive than hydroxyl radicals. The cell has much more time to dispose of them safely. Moreover, the superoxide radicals formed during respiration are produced at very particular locations within the cell, and we have evolved to deal with their normal production at these sites. There may also be a threshold of repair, linked to the rate of damage. In the course of normal respiration, where damage to DNA presumably accumulates slowly, virtually all damaged DNA will be repaired. Following serious radiation poisoning, when a massive amount of damage is produced very quickly, it is clearly not.
But despite all this, there is no qualitative difference between the kind of protection that is effective against radiation poisoning and that which protects against oxygen toxicity. This was understood by Gerschman and Gilbert, and other early workers in the field during the 1950s, who reported that several antioxidants helped to protect mice against the lethal effects of both x-irradiation and oxygen poisoning.
The point is driven home forcibly by the case of an unusual bacterium that is so resistant to radiation — 200 times more resistant than the common gut bacterium Escherichia coli, and perhaps 3000 times more
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resistant than we are — that the controversial astrophysicist Fred Hoyle proposed it must have evolved in outer space. Hoyle put forward this theory in the context of an argument for panspermia (meaning ‘seeds everywhere’) in his 1983 book The Intelligent Universe. Bacterial spores are so resistant to radiation that they can float about in space more or less unharmed, despite the intense cosmic radiation. It is therefore conceivable that life on Earth could have been seeded from space. Hoyle’s ideas were updated by the cosmologist Paul Davies, in his book The Fifth Miracle, who agreed that such impressive radiation tolerance made little sense unless life was forced through a radiation bottleneck at some stage in the past.
The tiny monster that Hoyle and Davies describe is a red-pigmented bacterium called Deinococcus radiodurans, one of a tightly knit family of six similarly resistant cousins. D. radiodurans itself is among the most radiation-resistant organisms yet found on Earth. It was originally detected as a contaminant of irradiated canned meat, and has since cropped up in weathered granite rocks in a virtually sterile Antarctic valley, in medical equipment sterilized by radiation, and in many more normal environments, such as room dust and animal faeces. Its resistance to ionizing radiation is coupled with a more general resistance to other types of physical stress, including ultraviolet radiation, hydrogen peroxide, heat, desiccation and a variety of toxins. Such an enviable suite of qualities makes D. radiodurans an ideal candidate for the bioremediation of sites contaminated with radiation and toxic chemicals. These possible commercial applications have stimulated interest in its DNA genome —
its total collection of genes — and in November 1999, Owen White and a large team, mostly from the Institute of Genomic Research in Rockville, Maryland, published the complete genomic DNA sequence, that is, the readout of its complete four-letter DNA code, in Science. We now have a much better idea of how it works.
The bacterium is a chimaera, a wonderful example of nature’s ability to cobble together a hotchpotch solution and give it every semblance of preconceived design. There is no magic trick, and no need to invoke a stellar origin. Almost all the DNA-repair mechanisms present in D. radiodurans are also present in other bacteria, although usually not all together.
The only one unique to D. radiodurans is an unusually efficient garbage-disposal system, which discards damaged molecular building blocks before they can be incorporated into DNA during DNA replication or
Oxygen Poisoning and X-Irradiation: A Mechanism in Common • 127
repair. The amazing ability of the bacterium to survive insult and injury stems from its capacity to hoard multiple copies of its own genes, as well as other useful genes it acquired by transfer from other bacteria.6 While most bacteria survive quite happily with a handful of protection devices, D. radiodurans uses the complete repertoire, and has multiple copies of each. This allows it to flourish in hostile environments, where it has little competition from less well-endowed brethren.
Far from the cosmic radiation bottleneck suggested by Davies, it seems likely that Deinococcus may have adapted relatively recently to radioactive environments. In their Science paper, White and his colleagues compared the genome sequence of D. radiodurans with the sequences of other bacteria, and argued that the closest living relative of the superbug is an extreme heat-loving bacterium by the name of Thermus thermophilus.
Of the 175 known T. thermophilus genes, 143 have close counterparts in D. radiodurans, suggesting that its toughness may have originated through the modification of systems that evolved originally to provide resistance to heat.
There is an important general point to make here, which we will return to later in the book. The genes that protect against radiation are not only the same as those that protect against oxygen toxicity, but are also the same as many of those that protect against other types of physical stress such as heat, infection, heavy metals or toxins. In people, the genes activated by radiation also protect against oxygen poisoning, malaria or lead poisoning. The reason for this cross-protection is that many different physical stresses all funnel in to a single common damage-causing process in the cell, so all can be withstood through common protective mechanisms.
Gene transfer can complicate attempts to define the evolutionary lineage of bacterial species.
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signal to the cell that it is under threat. Oxidative stress is therefore both a threat and a signal to resist threat. In the same way, the bombing of Pearl Harbor was both an act of aggression and a signal to the Americans to resist the Japanese threat.
The integration of protective mechanisms against oxidative stress raises the possibility that life might have evolved ways of dealing with oxygen toxicity long before there was any oxygen in the atmosphere —
ionizing radiation alone might do the trick. We have already concluded that rising oxygen levels did not bring about a mass extinction during the Precambrian or afterwards (Chapters 2–5). As oxygen is undoubtedly toxic, life might somehow have adapted to the threat in advance. Could it be that life adapted to radiation early on, and that this formed the basis of its response to other kinds of threat? If so, then Fred Hoyle and Paul Davies may have been right in one sense. Life was forced through a radiation bottleneck, but it happened on Earth, not in space, and it happened not recently, but 4 billion years ago.
This possibility is supported by the conditions discovered by the Viking lander on the surface of Mars in 1976. The lander biology instrument had been designed to perform three experiments to determine whether life was present in the soils of Mars. The results of these experiments were not sufficiently clear-cut to support a definitive answer, and arguments about the correct interpretation of the data continue today.
But one of the studies, the gas-exchange experiment, produced results that were, if not clear-cut, at least a total surprise. The experiment was intended to distinguish between gas emissions arising from microbial metabolism and those arising from purely chemical reactions. The instrument incubated Martian surface samples under dry, humid or wet conditions, and then measured any gases released. The samples were treated with a nutrient broth consisting of a mixture of organic compounds and inorganic salts, described by Gilbert Levin (one of the original Viking scientists and a steadfast proponent of life on Mars) as ‘chicken soup’. The experiment was conducted in two stages. First the lid was taken off the broth, to allow the water vapour escaping from it to humidify the soil in the box; then a little soup was sprinkled onto the soil, to promote metabolism by any organisms present.
To the amazement of the team, removing the lid from the broth was enough to produce a great burst of oxygen from the Martian soil — 130
times more than they had anticipated. The investigators toyed with the
Oxygen Poisoning and X-Irradiation: A Mechanism in Common • 129
idea that the broth might have stimulated photosynthesis, but the same reactions took place in the dark, and even after the samples were heated at 145°C for 3.5 hours to kill any microbes. If the samples were recharged with fresh nutrient following the initial burst of oxygen, however, no more oxygen was released, implying that the reactions had come to an end. Although these results did not strictly rule out life, the reactions were much better explained by chemistry than by biology. But if this was the case, the chemistry of the soil must have been potent, as even the simple expedient of adding water made it fizz. After some puzzlement, the Viking team eventually concluded that the soil samples must have contained superoxides and peroxides, generated by ultraviolet rays acting on the atmosphere or on the soil itself. This conjecture has been confirmed by other analyses of the chemical composition of the rocks.
What had happened? We can infer that hydroxyl radicals, hydrogen peroxide and superoxide radicals were formed over the aeons by ultraviolet rays splitting traces of water vapour in the Martian atmosphere or soil.
Without surface water, these intermediates would have reacted with iron and other minerals in the soil to produce the rusty oxides that give the red planet its colour. Most of these metal oxides would be unstable on the Earth, but on Mars they remained petrified, precariously stable, as long as the dry and sterilizing conditions persisted. When the Viking scientists took the lid off their broth, the suspended chemical reactions could finally run through to completion. As the unstable iron oxides broke down, the petrified intermediates reacted by way of the pathways we have discussed in this chapter to conjure up oxygen and water from the very rocks themselves. Ironically, the science-fiction heroes struggling to detoxify the soil and replenish the air of Mars with oxygen may need no more than a little warm water to turn the Red Planet blue.
We must conclude that Mars is under serious oxidative stress.
Although its thin atmosphere contains barely 0.15 per cent oxygen, any hypothetical life dwelling there would have to contend with oxygen toxicity generated by radiation, certainly as serious as anything faced by life on Earth today. If this is the case on Mars today, it must surely have been the case on the Earth 4 billion years ago. The Earth, after all, is closer to the Sun, and subject to more intense solar radiation. Before there was any oxygen, there was no ozone layer, and the cruel ultraviolet rays must have cut straight through to the ground. But the traditional view, that the continents and shallow seas would have been sterilized by the
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rays, is being turned on its head. New evidence suggests that resistance to oxygen and radiation alike was built into the very earliest organisms, rather than being built in later as an afterthought. The implications for evolution, and for our own make-up, run deep, as we shall see in the next few chapters.
C H A P T E R S E V E N
Green Planet
Radiation and the Evolution of Photosynthesis
The hitch-hiker’s guide to the galaxy describes the Earth as an utterly insignificant blue-green planet, orbiting a small and unre-garded yellow sun in the uncharted backwaters of the western spiral arm of the galaxy. In deriding our anthropocentric view of the Universe, Douglas Adams left the Earth with just one claim to fame: photosynthesis. Blue symbolizes the oceans of water, the raw material of photosynthesis; green is for chlorophyll, the marvellous transducer that converts light energy into chemical energy in plants; and our little yellow sun provides all the solar energy we could wish for, except perhaps in England. Whether Adams intended it or not, his scalpel was sharp —
photosynthesis defines our world. Without it, we would miss much more than the grass and trees. There would be no oxygen in the air, and without that, no land animals, no sex, no mind or consciousness; no gallivant-ing around the galaxy.
The world is so dominated by the green machinery of photosynthesis that it is easy to miss the wood for the trees — to overlook the conundrum at its heart. Photosynthesis uses light to split water, a trick that we have seen is neither easy nor safe: it amounts to the same thing as irradiation. A catalyst such as chlorophyll gives ordinary sunlight the destructive potency of x-rays. The waste product is oxygen, a toxic gas in its own right. So why split a molecule as robust as water to produce toxic waste if you can split
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something else much more easily, such as hydrogen sulphide or dissolved iron salts, to generate a less toxic product?
The immediate answer is easy. For living organisms, the pickings from water-splitting photosynthesis are mu
ch richer than those from hydrothermal activity, the major source of hydrogen sulphide and iron salts. Today, the total organic carbon production deriving from hydrothermal sources is estimated to be about 200 million tonnes (metric tons) each year. In contrast, the amount of carbon turned into sugars by photosynthesis by plants, algae and cyanobacteria is thought to be a million million tonnes a year — a 5000-fold difference. While volcanic activity was no doubt higher in the distant geological past, the invention of oxygenic (oxygen-producing) photosynthesis surely increased global organic productivity by two or three orders of magnitude. Once life had invented oxygenic photosynthesis, there was no looking back. But this is with hindsight. Darwinian selection, the driving force of evolution, notoriously has no foresight. The ultimate benefits of a particular adaptation are completely irrelevant if the interim steps confer no benefit. In the case of oxygenic photosynthesis, the intermediate steps require the evolution of powerful molecular machinery that can split water using the energy of sunlight. From a biological point of view, if you can split water, you can split anything. Such a powerful weapon must be caged in some way lest it run amok and attack other molecules in the cell. If, when the water-splitting device first evolved, it was not yet properly caged, as we might postulate for a blindly groping first step, then it is hard to see what advantage it could offer. And what of oxygen? Before cells could commit to oxygenic photosynthesis, they must have learnt to deal with its toxic waste, or they would surely have been killed, as modern anaerobes are today. But how could they adapt to oxygen if they were not yet producing it? An oxygen holocaust, followed by the emergence of a new world order, is the obvious answer; but we have seen that there is no geological evidence to favour such a catastrophic history (Chapters 3–5).