The Equations of Life

by Charles S. Cockell

  In more recent years, synthetic biologists, not content with changing the genetic code, have had great success in getting cells to incorporate new types of amino acids into proteins. With modern molecular tools, the hope is that some amino acids not used naturally in life might find use in proteins to make new therapies for diseases. Designer proteins, constructed from sets of amino acids not encountered in natural biochemistry, raise both enormous scientific potential and ethical questions.

  When looking at these newfangled creations, we might be tempted to use them as evidence that life is biochemically so flexible that the existing amino acids used by life must be just a fluke, a frozen accident. After all, if some of these new amino acids can accomplish new biochemical tricks, is it not the case that life has failed to tap into these capabilities, because to swap to the new amino acids would mean too great a disturbance to its existing pathways? Given the chance to rerun evolution, maybe life would find these new and exciting biochemical properties from scratch, building different sets of amino acids that would include those now in use by synthetic biologists?

  There is, however, a crucial difference between evolution and synthetic biologists. The scientists find particular biochemical properties that might be useful, perhaps to make an effective drug or a new compound of use to an industrial chemist or a pharmacologist. With forethought, the scientists can select an amino acid and incorporate it into the cell to achieve a desired result. Life, however, must select a set of amino acids that can be used across a vast number of proteins, and it must do this to optimize its energy demands. Having ten different sets of twenty amino acids, all of which might do something useful, costs a lot in terms of materials and energy. A cell that replicates sufficiently in the environment to proliferate with fewer energy-demanding pathways will likely be at an advantage. The same argument is true for expanded genetic codes. That we can add letters to the code at will and even make microbes with stable, larger alphabets in the laboratory does not demonstrate that over many millions of years, under exposure to natural environments and competition for food and resources, these expanded genetic codes would confer a long-term advantage to organisms against those with a code containing our familiar four letters.

  Philip and Freeland’s work illustrates that the pressure on life is more likely to result in a small generic tool kit of amino acids with a wide, even distribution of biochemical properties that maximizes the possible things that life can build with that set. This biochemical evolution differs greatly from the motivation and pressures that channel the energies of a synthetic biologist. The mere possibility that a cell can use an extravagant variety of amino acids to make proteins when coaxed by a scientist to do so tells us little about whether the selection pressures on a whole organism in the environment would preferentially land on those amino acids. The demand on life is rather to select the minimal number of maximally diverse types.

  We know that life can take diversions. The unusual amino acid selenocysteine is found in some proteins. The selenium atom within it seems to improve the ability of proteins to deal with antioxidants. Another strange cousin, pyrrolysine, is an amino acid found in some methane-producing microbes. Both compounds expand the set of amino acids in life to twenty-two, showing that when confronted with the need to expand the repertoire of protein building blocks, driven by some specific biochemical requirement, life can achieve this.

  The genetic code, with its number and types of bases; the codon table, which specifies the amino acids the table will code; and even the amino acids themselves are all apparently limited, nonrandom choices. However, perhaps all this does not matter. With just 20 possible amino acids, we have unlimited potential to string together a simply vast number of molecules. Consider a protein with 300 amino acids. Each place in the chain could have 1 of 20 amino acids. When strung together, these 300 places for any of 20 amino acids represent 2 × 10390 different possible combinations! That is enormously more than all the stars in the known universe. Therefore, with just a limited alphabet of amino acids, life has the unconstrained potential to create diversity, quirks, and experiments in design that knows no bounds. Here, surely, at the end of the trail, as the chain of amino acids finally folds up to produce a molecule, we are in the realms of chance. With such diversity, surely now we leave the constricting behavior of physical limits and open up a world of molecules where life in all its variety will be unbounded?

  When biochemists first began to explore the mesmerizing variety of proteins from which life is assembled, they seemed confronted by a paralyzing number to grapple with. With 2 × 10390 possible different sequences from a single hypothetical protein chain of just 300 amino acids, how many centuries of biochemical work would be necessary to make sense of all the molecules that exist in the real world? Yet as these molecules were uncoiled, their strings of amino acids read and their folds studied, it became apparent that no matter what the sequence of amino acids, the number of folds or shapes that parts of proteins could adopt was very limited indeed.

  Pull proteins apart into their individual units, and you will find that they parade a very meager set of folding arrangements. Helices (termed α-helices) are right-handed helical arrangements of amino acids held together by a hydrogen bond between the hydrogen on an amino group and an oxygen of an amino acid three or four places earlier in the sequence. Another type of fold are the pleated sheets (usually called β-sheets). These are long chains of amino acids held together by hydrogen bonds to make sheetlike arrangements.

  These two types of folds can be strung together to make combinations. Many proteins are made up of α and β structures, assembled from helices and sheets that occur in various permutations along the amino acid chain. In some proteins, the two forms occur in strictly alternating forms (α/β). These structures are themselves subclassified into triosephosphate isomerase barrel, sandwich, and roll motifs, which are particular ways in which the helices and sheets can be folded. Never mind the finer points; what we see here is a bounded set of possibilities.

  One explanation for this small chorus could be that these folds became locked into life early in its evolution and that, being sufficient to assemble something that is useful, there is no longer a selection pressure to evolve further forms. The analogy is like building a house. You do not go to a builder’s store and use every brand of brick available. You select a few that will do the job. Once an ancestral organism made the choice of folds, the rest of life was stuck with it.

  Compelling though that argument may seem, much more fundamental laws at work may select for the arrangements of protein folds. Amino acid chains will collapse in such a way as to arrive at a low-energy state. The successive folding steps are driven by thermodynamics to their most stable state. The folds are not independent, and different parts of a protein exert an influence on the folding patterns of other parts. As all these folds collapse into the final product, the protein seeks the most thermodynamically favorable configuration. This means there are only a few solutions. Are the laws of physics disobeyed, or entropy violated, if a strand of disordered amino acids neatly packages up into a ordered machine ready to do some work? Not at all. When amino acids arrange themselves into a protein, water molecules are being forced out of the structure into the surrounding milieu, into the disordered chaos of water molecules in the outside environment. The second law of thermodynamics is not violated in this transition to molecular order.

  Here again we see a beautiful synergy between biology and physics, sometimes instead framed as a polarized difference. Some people perceive a conflict between two possible views, between the existence of biological “laws” that drive life to a few simple and predictable solutions and a different, “Darwinian” perspective of evolution, where there is no preordained order, and variation and selection define a vast landscape of possibilities. However, the two viewpoints seem compatible and inseparable. Darwinian evolution, through genetic variation and selection, experiments with a great diversity of forms, but those forms still conf
orm to the laws of physics and are tightly constrained by the universal principles that operate at whatever scale we are observing. With proteins, Darwinian evolution generates many proteins with diverse structures useful for different functions, selected because the processes in which they are embedded are beneficial for survival. However, thermodynamics greatly restricts the number of shapes in which this glut of molecular forms can be assembled.

  The study of the genetic code and its translation into the fabric of living things has occupied many people. Some have a fascination for DNA, others for protein, still others for that long-lost Narnia of early Earth, where the first RNA molecules may have leaped into chemical activity and life. A few spread themselves across this landscape of biochemistry. However, over the last few decades, independently across this vista, scientists have apparently removed contingency from much of what was once considered a virtual miracle of machinery. Life was believed to be a system of such extraordinary molecular complexity and yet, in its functioning, a system of such elegant simplicity, it seemed the whole thing must have been chance, an accident, one of many paths that life might have taken. Yet physical and chemical constraints appear to have hammered and forged the code of life in ways that now, with computational methods, even open themselves to comparison with alternative worlds. The fog lifting over this once-unnavigable diorama of molecular forms reveals it instead to be aligned and arranged in patterns more clearly seen.

  CHAPTER 8

  OF SANDWICHES AND SULFUR

  WHEN I UNWRAP ANOTHER sandwich at the café in the building in which I work in Edinburgh, it doesn’t usually consciously occur to me that this culinary delight is not a mere sandwich, but a bundled-up package of tasty electrons, subatomic particles that come in lettuce, tomato, and chicken flavor.

  Yet, hidden behind its sandwichy guile, this tempting cardboard-encased cocoon of university catering sumptuousness is nothing more than a convenient way of consuming electrons. Throughout the vast diversity of living things, from the smallest bacterium to a blue whale, there is a stunning commonality in how the cells of these creatures get their energy to grow and reproduce. So identical is this process across life, so simple and in which such basic principles inhere, that it is easy to imagine that life anywhere across the universe might get its energy in the same way. It is in this machinery that we pursue our adventure into the structure of life, exploring its basis in physical processes. From the codes and molecules that build life, we turn to another vital piece of life’s molecular machinery—how it gathers energy from its environment to grow and reproduce, the process that powers the biosphere.

  In the 1960s, a brilliant scientist, Peter Mitchell, pondered the basic mechanisms of how life gathers energy from the environment. He wondered about this because he knew the question was important. The second law of thermodynamics that drives the universe inexorably toward disorder or increasing entropy is a fact of the universe; hence, it is a law, and life must conform to it. Constructing complex machines that can grow and reproduce requires energy to maintain this order against the ever-present second law, which would like to dismantle those machines and dissipate their energy and the components of their molecules into the void. Fathoming how life gets energy from its environment is therefore not merely of interest to apprehending how it interacts with the world around it, but it is fundamental to knowing how it operates within the constraints of the laws of the universe; few are as basic as the second law. In the temporary oasis that is our Earth, receiving energy from the Sun and producing its own from the primordial heat within the planet, how does life gather its energy to garner such organizational complexity and then to spread tenaciously across and within our planet?

  Mitchell’s biochemical insights eventually won him the Nobel Prize in 1978. Like many findings that reshape our worldview, they seem obvious with the benefit of hindsight, but putting the bits together in a way that, to later generations, would appear to be common sense required a stroke of creative genius. The result was yet another foundation stone in our understanding of biology—an understanding that speaks of general applicability, another basic piece of machinery whose roots in physics suggest the potential similarity of life across the cosmos, if indeed it has evolved elsewhere.

  But back to the sandwiches. Once consumed, where do those sandwiches go? They are disassembled in our bodies into their constituent sugars, proteins, and fats. Some of that material is burned in oxygen to release energy, while some of it may be indigestible. Now in secondary school, you may well remember being tirelessly forced to write down the reaction for aerobic respiration, and dull it was. But bear with me, because one thing they never taught you at school was just how beautiful this process is.

  C6H12O6 + 6O2 → 6CO2 + 6H2O + energy

  There on the left we have a complex chemical formula, C6H12O6, which is the formula for glucose, a sugar, but it could be any complex carbon compound, from the ingredients of sandwiches to salami. We add this carbon compound to oxygen in the air we breathe, and when we do, these two compounds make energy, with carbon dioxide (CO2) and water (H2O) as waste products on the right-hand side of the equation.

  The organic material, the sugar shown above, contains electrons, each in a fuzzy orbit within its atom. The electrons contain energy, and it is this energy that life seizes from the reaction. But how does it do that?

  All atoms in the universe have varying degrees to which they will give up their electrons. Many of these atoms are electron donors, relinquishing electrons with glee, but others, electron acceptors, prefer instead to take them up. Whether an atom donates or accepts electrons is influenced by a concatenation of things, from pressure and temperature to acidity, but we need not concern ourselves with this detail here. Crucial to your lunchtime hunger is that many organic materials, including the components of your sandwiches, are good electron donors.

  Sitting on cell membranes or the membranes of the organelles within them, for example the mitochondria in your cells, there are molecules that will bind to compounds with electrons ready to be grasped. The electrons have now begun the first stage of their journey. Through an act reminiscent of a relay race, the electrons are passed from the broken-down products of your sandwiches to the cell.

  Sitting next to the molecule that has just grabbed the electron is yet another molecule that would like the electron even more, and so the particle begins its traverse through the cell membrane, leaping from one molecule to the next. The relay race is under way. Eventually, the electron will get to the end of the race, and what then? There sits an electron acceptor that would like to grab the electron and carry it off. In our bodies, that is the job of oxygen. The electron acceptor is crucial because if you do not take the electrons away, they will get clogged up in the cell and we quickly get an electron traffic jam. The transfer process we have just been talking about will grind to a halt. And now you can understand why breathing is important: getting that oxygen into your body to stop your energy machinery from overloading is rather crucial.

  As the electrons shunt through the membrane, their energy is released and we now must do something with that energy: we must gather it up. Mitchell creatively figured out how this was done. As each molecule gains that minuscule amount of energy from the electron as it passes by, the molecule uses the energy to move yet another subatomic particle, a proton, from the inside of the membrane to the outside.

  What we now have is a proton gradient: more protons are on the outside of the membrane than in the inside. Those protons, through osmosis, now want to move back into the cell to equalize the gradient.

  Place a raisin in a cup of tap water, and the fruit will expand as the salty and sugary interior sucks up water from its surroundings. This is osmosis at work. The water will move in a direction where it can end up with equal concentration inside and outside the raisin. In the same way, as we now have more protons on the outside of the membrane than on the inside, the interior will slop them up until the concentration on both sides of the cell membr
ane is the same.

  The protons sitting outside the membrane have a Janus-faced quality: not only are they at a higher concentration, but they also have a positive charge to them, each one written as H+. It is both this higher concentration of charge on the outside of the membrane, designated ΔΨ, and the higher concentration of actual protons themselves, written as ΔpH, that creates this powerful gradient. We call this gradient the rather dynamic-sounding proton motive force, Δp, and the equation to work it out is below:

  Δp = ΔΨ − (2.3RT/F)(ΔpH)

  where R is the universal gas constant (8.314 J/mol/K), T is the temperature of our cell, and F is the Faraday constant (96.48 kJ/V). A typical value for the proton motive force is about 150 to 200 millivolts.

  So the protons have a tendency to be drawn into the cell. This they now do, but not by randomly diffusing through the membrane anywhere they please, since the membrane is generally impermeable to them. They flow back through a complex little machine called adenosine triphosphate (ATP) synthase, whose job it is to make the energy-trapping ATP.

  As the protons flow back through ATP synthase, they cause the pieces of the molecular machine to rotate. This incredible contraption is made of no less than six different protein units. The changing shape of ATP synthase as it goes on its ratchet roundabout brings phosphate groups into alignment with adenosine diphosphate (ADP) and forces them together to make ATP. These new phosphate bonds have now trapped the energy of the electron transport chain.