Science in Christian Perspective

The Formation Of Living Organisms From Non-Living Systems*

WALTER R. HEARN

Assistant Professor of Chemistry, Iowa State College

From JASA 10 (December 1958): 3-8.

In 18-59, Charles Darwin cautiously stated, "Science as vet throws no light on the ... problem of the essence or origin of life."¹ Actually, the possibility of observing the "spontaneous generation" of living things from inorganic matter was at that time a very controversial scientific question, as Pasteur had not yet completed his conclusive experiments along these lines². However, instead of praising Darwin for this reluctance to extend his theory too far, critics of Darwin and of the theory of natural selection have often attacked at this very point, saying that the power of the theory to account for biological variations is vitiated by its failure to account for biological origins. Thus Clark, in 1950. stated that the influence of the theory of natural selection probably did biology more harm than good, by concentrating attention on small changes in living things and diverting it from the problem of their origin. He went on to say:

"For the materialist, organisms are limited by the properties of atoms and molecules. Somehow or other, he must explain how chemical molecules of gigantic complexity came into existence and have been able to arrange themselves in increasingly complicated ways. This is the fundamental problern of evolution, yet it is generally ignored in modern books on the subject, nor, (at least to the author's knowledge) has the problem ever been fairly faced ."³

Oparin's book on the origin of life4 was published in English translation in 1938 and certainly faced the problem fairly. The current scientific literature abounds with papers facing the problem, and today there is no reluctance at all to extend Darwin's principle of natural selection into the pre-biological realm, making the origin of living organisms merely a consequence of a still more fundamental "chemical evolution" of non-living molecules. The tendency today is to regard the original spontaneous development of living things from inanimate matter as not only highly probable, but actually inevitable5. What are the factors which have led to this recent extension of the evolutionary concept?

In the first place, the time available for a process of chemical evolution is now generally accepted to be

* Paper presented at the Twelfth Annual Convention of the American, Scientific Affiliation, Beverly Farm&, Massachusetts, August, 1957.

about two billion years⁶, which is regarded as adequate if the process I s considered to be the over-all result of a large number of continuing small changes. Since Oparin4, this gradualism has been a predominant feature of the theory, and has largely served to discredit arguments such as that of Leconite du Nouy⁷, who calculated that the formation of a single molecule of protein would require 10²⁴³ billions of years, if the probability of the chance collision of its constituent atoms were the only factor taken into consideration. If a series of stepwise reactions is postulated rather than a single all-or-none jump from simple molecules to complex ones, the probabilities dealt with are entirely different, and such calculations become meaningless.

A second factor, I think, has been an increased appreciation for the stability of biologically important molecules. Two previous papers in this symposium have described experiments in which mixtures of very simple compounds have been submitted to conditions simulating pre-biological conditions on the earth, and have produced more complex molecules identical to those common to living organisms today^8,9. Undoubtedly such experiments demonstrate the intrinsic stability of amino acids and other such compounds and made their prebiological accumulation on the earth's surface highly probable. Another line of investigation is that of Abelson and others in the field of paleobiochernistrylO. It has been found that recoverable amounts of the original organic matter are present in such fossil materials as shells ' bones, petroleum, and coal. and possibly even in the Pre-Cambrian black shale. For example, it was possible to study the content of protein and amino acids in the shells of clams (Merccizaria mercenaria, the quahog of the Atlantic coast,) now living and in those of the same species 1,000, 500,000, and 25 million years old. The 1,000 year old specimen had the same protein content and the same amino acid pattern as the modem specimen; the 500,000 year old specimen contained no protein and smaller amounts of amino acids, some of which were still in peptide chains; and the 25 million year old shell contained only a few of the original amino acids and no protein or even peptides. Laboratory experiments on the stability of alanine at various temperatures indicated that this amino acid would be sufficiently stable at a temperature of 20deg C for half of the original amount present to remain after a period of about three billion years. Substantial quantities of porphyrins, compounds of the same type as the chlorophylls or the heme of important respiratory pigments, have been found in petroleum samples at least tens of millions of years old. Such studies not only open up the exciting possibility of discovering something about the metabolism of early forms of life, but also emphasize in a dramatic way the inherent stability of these biologically important compounds. If their pre-biological synthesis was possible, as I think has been more or less clear]-, demonstrated, then they must have accumulated and set the stage for further developments.

Perhaps it should be pointed out here that our notions of the instability of large complex molecules rest partly on theoretical grounds and partly on the grounds of common experience. If chemical bonds have a certain probability of being broken, it stands to reason that under the same conditions a molecule with a large number of bonds will more likely be broken down to smaller molecules than will a molecule which has fewer susceptible bonds. However, organic chemists know that chemical bonds in a molecule may affect each other in different ways: in some cases a bond is "labilized" by the presence of other bonds (that is, made more susceptible to attack), but it is also possible for bonds to be "stabilized" by the influence of other bonds (for example, as by resonance in the case of the porphyrins already mentioned, or by the formation of zwitterionic dipoles in the case of the amino acids). In the case of very large molecules, such as the proteins and nucleic acids, steric factors as well come into the picture. Thus, with a molecule of sufficient size, forces such as those due to van der Waals attraction or hydrogen bonding may become intra-molecular, and their combined strength may impart considerable stability to a particular configuration of the molecule. Pauling and others have demonstrated this in the case of helical structures for certain proteins", and the WatsonCrick double helical structure for DNA (desoxyribose nucleic acid) is an even more striking example12.

It is not so much these theoretical considerations, therefore, as it is our practical experience which leads us to believe intuitively in the instability of complex molecules. The phenomena of decay and decomposition are such commonplace experiences with us that I think we often fail to realize how much of this is actually biological. The earth's surface and atmosphere today is literally teeming with living organisms whose existence depends on their ability to degrade complex molecules and derive energy thereby. In Pasteur's day, people found it almost incredible that the air they breathed should be swarming with microorganisms and their sporeS2, and sometimes even today we forget that we live in such a biologically crowded atmosphere. If we lived tinder circumstances in which food did not spoil at ordinary temperatures and mold did not grow on our protein solutions in the laboratory, we might not have such a strong feeling that complex molecules are intrinsically unstable. But, if the theory of chemical evolution is correct, complex organic compounds were synthesized and did accumulate in just such a "sterile" or "a-biological" atmosphere.

In general terms, what is it that we are seeking to understand about life in terms of the laws of chemistry and physics? It may seem strange that it is actually quite difficult even to define "life" or "living" satisfactorily. This difficulty has been emphasized by Pirie¹⁵, who pointed out that anv definition of life which would exclude all obviously non-living systems would probably also exclude some obviously living systems, and vice-versa. Pirie has set tip what he considers generally acceptable criteria for a system to be regarded as "living" as a sort of ground-rule in lieu of a precise definition, in case someone claims to have produced such a system artificially ¹⁶. If we cannot say precisely what a living thing is, we can at least specify some of the things it does. We might say in the most general terms that living systems are systems which are able to utilize a flow of energy through the system to maintain a higher degree of molecular order than their surroundings (or in thermodynamic terms, to Ctextract" negative entropy from the energy sources utilized by the system). In addition, although a system without the ability to reproduce may be regarded as living (a castrated animal, for example), at least the Possibility of self -duplication must be present under certain conditions for a system to have much significance in evolutionary considerations. We should also keep in mind that if the Darwinian idea of variations plus natural selection is applicable to the pre-biological realm, variant systems of metabolism may have arisen and become "extinct" just as variant morphological  forms seem to have done in the biological realm. In duction system to another, each of higher oxidation fact, the possibility exists of discovering such variant potential, until finally two hydrogens combine with an systems, possibly even as "vestigial remains" in modern atom of oxygen. There are some strictly anaerobic organisms.

forms seem to have done in the biological realm. In fact, the possibility exists of discovering such variant systems, possibly even as "vestigial remains" in modern organisms.

The standard procedure for studying any complex system is to resolve it into simpler components which can be studied separately, or perhaps to construct a model of similar but less complicated design and to study the model first before attempting the more difficult study of the system itself. Thus, we might resolve living things into systems for obtaining energy and systems for utilizing this energy (or the entropy deficit accompanying it) for synthetic purposes; in addition we might investigate the way these two systems are "coupled," and ways in which the entire complex is held together or reduplicated. In practice this is what biochemistry has been doing in recent years -, with varying degrees of success in elucidating the details of the different systems. It must be admitted that we do not as yet have a complete picture of any of the isolated systems, but we do have a partial picture of each of them. What is most significant here is the fact that it has been possible to study such isolated systems in this way at all. Now, let me sketch in a broad outline of those metabolic processes which may be said to have had the -reatest "survival value." and then we shall look at some possible ways in which they might have developed from less complex processes.

There seem to be three major processes by which living things obtain energy. These are oxidation, fermentation, and photosynthesis. Actually, if we limit ourselves for the moment to energy-yielding processes, the latter should really be called "photodecomposition," since the synthetic part of the process actually utilizes the energy thus obtained.

Oxidative metabolism makes use of molecular oxy en to give final products of carbon dioxide and water in the usual scheme in which organic compounds are oxidized. In a series of steps, hydrogen atoms are removed from the substrate, with or without a preliminary addition of a molecule of water, and eventually made to react with oxygen. The carbon dioxide is liberated when the substrate has been rearranged to form a carboxylic acid with unsaturation on the beta-carbon atom, a type of compound which readily decarboxylates. To oxidize a two-carbon compound such as acetic acid, the compound is first condensed to a "carrier" molecule in order to have enough carbon atoms to form the beta-keto acid analog; in the course of the subsequent oxidations and decarboxylations, the carrier molecule is regenerated, giving a cyclic process. The energy-yielding part of such a cycle, the Krebs citric acid for example, is the oxidation of the hydrogen to water. In all cases of oxidative- metabolism investigated, this is a stepwise process in which the hydrogen atom, or an electron accompanied by a hydrogen ion, is transferred from one oxidation-reduction system to another, each of higher oxidation potential, until finally two hydrogens combine with an atom of oxygen. There are some strictly anaerobic microorganisms which apparently do not make use of this process at all, and there are also autotrophic chemosynthetic bacteria which oxidize sulfur, iron, or hydrogen principally rather than carbon cornpounds¹⁷.

Fermentation, an anaerobic process, consists basically of the oxidation of one compound by reducing another compound other than oxygen, so that the presence of oxygen is not required. In some cases, this oxidation-reduction may be intra-molectilar, as in the dismutation of glucose to lactic acid in the familiar Embden-Meyerhof scheme. In this case, which also involves a cleavage into two smaller molecules, it might be said that one end of the molecule has been oxidized (the carboxyl group of the lactic acid) while the other end has been reduced (the methyl group). Anaerobic processes are often found in tissues whose priniarv mechanism for obtainintg energy is oxidative, as mammalian muscle.

Of the mechanisms involved in coupling these synthetic processes to the exergonic ones, we have varying degrees of knowledge. We have come to realize that the only possible way such coupling can take place is through the raising of a specific chemical compound to a higher energy state by the rearranging of the electronic configuration of its constituent atoms. 

In some cases we know what the compound is, and how the rearrangement takes place. In many cases there is an intermediate compound which can serve as an energy "carrier" because it is thermodynamically unstable but kinetically stable; such a compound transfers its energy only when a suitable catalyst is present to speed tip the reaction. Perhaps the most common example is that of ATP (adenosine triphosphate), which has at one end of the molecule two pyrophosphate groups which show this property of thermodynamic instability (i.e., "high energy") but which are not hydrolyzed at an appreciable rate at the ordinary temperatures and pH's of living cells and tissues. When a suitable catalyst is present. ATP is rapidly hydrolyzed, and we have a rather definite picture of how the catalyst accomplishes this²⁰. ATP seems to play a role in all types of metabolic processes, and in some cases we know exactly how it works. For example, in the anaerobic fermentation of glucose we know exactly how the energy is used to synthesize ATP, and exactly how many molecules are synthesized. In the case of oxidative metabolism, we know that ATP is synthesized, and we know approximately how many molecules must be synthesized for every hydrogen atom that eventually combines with oxygen, but we do not know in this case the actual chemical details of the synthesis²¹.

As for mechanisms by which metabolic interrelationships are preserved and duplicated in reproduction, it must be said that we have probably just discovered the most important clues but have not yet had time to solve the mystery. The identification of the molecule that actually carries the hereditary pattern as DNA²², and the elucidation of the major features of its structure¹² have certainly given its hope for the first time of understanding heredity at the molecular level. It is interesting to note that DNA is made up entirely of molecules of the same general type as ATP minus the high-energy pyrophosphate, and that the structural features of the molecule already account beautifully for many of the properties of the hereditary carrier, such as stability in thread-like form and ease of exact duplication.

Now, extrapolating backwards, what can we say about the sequence of events leading to the accumulation of these systems into what we know as living things? In the first place, if the original atmosphere was really a reducing one rather than an oxidizing one, it is highly probable that oxidative metabolism was not the original exergonic process utilized. A number of authors have regarded anaerobic fermentation as the most likely original pathway; Kirkwood points out that it may be regarded as possibly more "primitive" than oxidation or photosynthesis because the anaerobic enzymes are usually soluble, while the others are usually intimately connected with cell structure²³. If the atmosphere gradually became more oxidizing, due to the photolysis of atmospheric water and the loss of the lighter hydrogen resulting, then oxidative metabolism may have developed. Oxidation is capable of yielding much more energy than fermentation; with the large quantity of organic compounds available frorn prebiological synthesis, this would have been a very advantageous scheme-until, of course, the supply of organic compounds ran out! The oxidation of iron and sulfur may have been resorted to, but such schemes would also be heading up a blind alley. There may have occurred crisis after crisis, but eventually the atmosphere would have become too oxidizing to replenish the supply of reduced molecules; some other scheme must have developed, in this case, photosynthesis. The energy of sunlight could then be used to reduce carbon dioxide resulting from oxidative metabolism, and the familiar carbon cycle of nature as we know it today was underway.

Is it reasonable that metabolizing systems should arise spontaneously and persist? The systems we know today are much too complicated to have appeared suddenly, but there is yet the possibility that the system might have evolved by the continued modification of only slightly advantageous chemical reactions. Several suggestions have been made along these lines. In a general sense, any chemical reaction which produced a compound with even slight catalytic power to stimulate the reaction by which it was produced would have

survival value" over competing reactions. Calvin6 has pointed out that iron-porphyrins are catalysts which will promote certain steps in the synthesis of porphNrins from succinic acid and glycine, two contpounds alreadv known to be synthesized tinder model "prebiological" conditions; furthermore, since iron itself has very slight catalytic power to do the same thing, we have a good example of a synthetic reaction which would be expected to "evolve" from very simple precursors. It might be mentioned that the further combination of the iron-porphyrin compound with a specific protein increases its catalytic power many thousandfold, but this elaborate refinement probably came much later in the development of this system.

It has also been pointed out that inorganic pyrophosphate itself has the same properties of thermodynamic instability plus kinetic stability as ATP, which may be regarded merely as pyrophosphate with an organic "handle" on it. Therefore, pyrophosphate could have served as one of. the first molecules capable of transferring energy from exergonic reactions to endergonic synthetic reactions. Kirkwood²³ suggests that pyrophosphate probably accumulated from thermal reactions along with the pre-biological supply of organic compounds, and Calvin⁶ has even suggested a very rudimentary coupling reaction for the synthesis of pyrophospbate from orthophosphate, linking the dehydration of orthophosphate to the squeezing out of a molecule of water from the coordination sphere of  iron or vanadium ions are oxidized. It is thus possible to devise fairly reasonable models for "primitive" metabolic schemes, a stimulating hobby of growing popularity among biochemists.

We do know from recent studies on enzymes and co-enzymes that simple molecules may often have enough catalytic power without combination with a specific protein to make such primitive metabolizing systems seem reasonable. Some attempt., have already been made to design a "mathematical model- of the evolution of primitive metabolizing systems by making various assumptions about possible effects the products might have on the catalysis of their svnthesis²⁴. For example, a compound would not have to serve as a specific catalyst in order to promote its own synthesis over that of other compounds; if it were a macromolecule it might serve just as well by adsorbing the reacting molecules or even by forming a prototype of a cell wall to hold the system together. It is probably too soon to assess the validity of the various mechanisms proposed^{5,23,25,26,27} but the general principle does seem valid in the light of our present knowledge of biochei-nistry²⁸. The difficulty of obtaining conclusive data will no doubt produce a great deal of speculation. such as the argument over whether the assembling of metabolizing systems most probably began by adsorption on inorganic clays, as proposed by BernaI²⁹, by aggregation in quiet waters of ocean depths, as proposed by Pringle³⁰, or in water-logged soil or layers adsorbed at air-water interfaces, as proposed by Haldane^3l. However, the concept of pre-biological evolution results not only in speculation but also in the stimulation of specific lines of research³², such as the investigation of "primitive" types of organisms, of variant pathways of metabolism, and of model enzyme systems.

Continuing biochemical research on "sub-vital" systems may be expected to lead to further explanation of biological mechanisms at the molecular level. Further research on cell nuclei^33,34, on crystalline DNA which "transforms" bacteria to other genetic types^35,36, on bacteriophage which inject DNA into bacteria and bring about replication of the phage^37,38, on TMV (tobacco mosaic virus) protein and RNA (ribose nucleic acid) which recombine to form an infective virus particle ^39,40, on the "electron transport particle" of mitochondria^4l,42, on chloroplasts which photosynthesize ^43,44, on protoplasts which synthesize induced enzymes⁴⁵, and so on, will probably intensify the conviction that the living cell is an already highly developed system, a milestone perhaps, but not the starting point, of evolutionary processes.

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