Prebiotic evolution: Conformational perspectives

J. theor. Biol. (1982) 96,21-38

Prebiotic

Evolution:

Conformational

ANTONY

Perspectives?

W. BURGESS

Tumour Biology Branch, Ludwig Institute for Cancer Research, P.O. Royal Melbourne Hospital, 3050, Victoria, Australia (Received

28 October

1980, and in final form 22 October 1981)

Current models of prebiotic evolution are based on the chemical properties of proteins and nucleic acids. Even if it is conceded that these polymers can be synthesized abiotically, it is conceptually difficult to construct a link between them which would serve as a primordial model from which the present day genomecould have evolved. The abundance of monosaccharidesin biological systemsand the conformational propertiesof polysaccharidessuggestan alternative model for prebiotic evolution. This model discardsthe principle of retrocontinuity in order to develop the connection between a primordial genome and the self-ordering DNAprotein synthetic system of today. In essence,the model proposesthe developmentof a self-orderingpolysaccharidemolecularsystemin which the positionsof atoms or groupsof atoms in spaceoccur repetitively as a result of the conformational preference of the polymer units (i.e. the moleculesare spatially ordered). It is proposed that ordered polysaccharidesformed within micellar environmentswhichthen grew to a critical size before splitting to restart the cycle. Catalytic polysaccharidesand glycolipids shouldbe capableof generatingsucha system.The conformational, catalytic and solution propertiesof thesemolecules,together with their likely abundancein prebiotic timesallowsthe constructionof a model self-orderingsystemcapableof evolving towardslife aswe know it. Introduction Prebiotic evolution has been the subject of considerable intellectual and laboratory investigation for many centuries (Leukippos, 460 B.C.; ValleryRadot, 1922; Eigen, 1971). These approaches have been dominated not

only by the state of knowledge of the molecular basis of life but also the fashionable views on the essence of life (Schrodinger, 1944; Crick, 1970). Current experiments and theoretical models of evolution appear to be biased towards the generation of nucleic acids and proteins (Crick, 1968; Nelsestuen, 1979; FOX & Dose, 1972; Eigen et al., 1981) and their molecular interactions. But is this where evolution started? t This work was in part performed at the Walter and Eliza Hall Institute for Medical Research at which time it was supported by the National Health and Medical Research Council of Australia. 21

oozz-5193/82/090021+15

$03.00/0

@ 1982 Academic Press Inc. (London) Ltd.

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The principle theme of our current thinking is illustrated in a statement by Dr. Leslie Orgel: “it is very difficult to see how a totally different biological organization could have undergone a continuous transition to the nucleic acid-protein system with which we are familiar” (Orgel, 1968). Consequently most prebiotic experiments have been designed to simulate the synthesis of “proteins” or nucleic acids (Fox & Yuyama, 1963; Armstrong et al., 1977), and most models for prebiotic evolution assume that the initial self-ordering events involved these two macromolecules (Eigen et al., 1981; Crick 1968). Yet, neither proteins nor nucleic acids appear to have the conformational or chemical properties which would independently facilitate the initiation of a “life-like” system, i.e. the development of a simple self-ordering cycle. Further, even accepting the unlikely possibility that one or the other of these polymers formed the basis of a self-ordering primordial cycle, no satisfactory molecular mechanism has been put forward to explain how the transfer of information between proteins and nucleic acids could have arisen. Currently there is a revolution in molecular biology which has changed our understanding of gene structure and its regulation (Robertson, 1978; Gilbert, 1978). The existence of DNA insertion elements, cutting and splicing enzymes have already been related to the generation of diversity and the evolution of particular genes (Gilbert, 1978; Doolittle, 1978; Reanney, 1979). However, there are further ramifications arising from the existence of introns which may reflect the origin of the present genome. In particular, the fact that many eukaryotic genes are discontinuous, raises the possibility that the initial self-ordering events need not have arisen from polymers which contained linearly contiguous information, but instead may have developed from informational domains separated on the surface of a molecular template. Evolution from polysaccharides would lead naturally to the formation of such a dispersed genome. Thus, the primordial polysaccharide hypothesis developed below, deliberately relaxes Orgel’s principle of retrocontinuity, preferring instead to explore the generation of a chemically feasible selfordering molecular system, which could subsequently evolve towards the present day protein-nucleic acid system. This initial system is based on the structural fidelity and catalytic activity of polysaccharides as well as the conformational and solution properties of monosaccharides and glycolipids. Problems

with Proteins

and Nucleic Acids

Three difficulties arise immediately when one attempts to impute initial prebiotic cycles to nucleic acids or proteins: firstly, the conformational

PREBIOTIC

EVOLUTION

23

properties of protenoids do not lend themselves to the formation of a self-replicating polymer; secondly, polynucleotides relevant to information storage have not been generated in abiotic experimental systems; and thirdly, the present day linkage between nucleic acids and protein biosynthesis involves an intricate recognition system which is conceptually difficult to reduce to a chemically feasible abiotic self-ordering system. Our present day enzyme system for the synthesis of proteins (e.g. the amino acyl transferases) requires the simultaneous recognition of a particular amino acid and its specific nucleotide triplet (copied from genetic information). This level of sophistication must be long removed, in time, from the initial events leading to the evolution of self-ordering molecular cycles. Indeed, it is at the transition point between nucleic acids and proteins where our current theories of prebiotic evolution falter (Reanney, 1979,1977). Whilst the abiotic synthesis of nucleic acids and proteins is an acceptable proposal, no plausible mechanism has been produced by which either polymer could become self-replicatory. A system based on the generation of an enzymatically active protenoid capable of catalyzing the formation of a nucleic acid which could induce the synthesis of similar protenoids is unsatisfactory. Protenoids do not have conformational integrity. The conformational space available to each residue (Fig. 1) allows a huge variety of three-dimensional structures to be generated (Anfinsen & Scheraga, 1975). Where the amino acid sequence is likely to fluctuate, the repeated generation of a particular threedimensional structure with self-ordering properties would be extremely unlikely. In the absence of a guiding template, the concentration of a particular catalytic protenoid, could not be expected to increase significantly. At best random coil polypeptides with asymmetric charge distributions may have been generated. Arguments have been made that negatively charged polypeptides with only glycine, alanine, aspartic acid and valine may have been the initial polymers (Eigen et al., 1981). Random sequence copolymers of these amino acids would only be able to act as “molecular glue” -where the conformational properties would not be so critical. Such polymers would have been ideal for facilitating aggregation of polysaccharide modified with the nucleic acid bases. Similarly, nucleic acids also have considerable conformational flexibility (Shindo et al., 1979). Although, it is possible for sophisticated versions of these polymers to form compact structures with unique conformation (e.g. tRNA, Sussman & Kim, 1976), recent measurements using nuclear magnetic resonance spectroscopy (Hogan & Jardelzky, 1979) and picosecond time dependent fluorescence depolarization (Millar et al., 1980) indicate that random sequence single stranded DNA and RNA have considerable

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(a)

Ib)

eqLdP 120

Y

0

-120

-120

0

120

a

FIG. 1. Conformational energy surfaces for units which determine the local conformations and flexibility of protein (a, b) and polysaccharide chains (c). (a) The simplest amino acid (glycine) has considerable rotational flexibility on either side of the peptide bond, approximately 50% of the available “glycyl” peptide conformations (-) are within 5 kcal/mole of the energy minima (Anfinsen & Scheraga, 1975). (b) Alanine and other amino acid residues with side chains, direct the folding of the protein chain more definitively (approximately 15% of the conformations for the.bonds on either side of the peptide unit, 4 and $I), are within 5 kcal/mole of the most favorable structure for the alanyl peptide (Anfinsen & Scheraga, 1975). (c) Rotation around the 1 + 4 p glycosidic is considerably more restricted even in simple carbohydrate polymers such as cellobiose. Conformational analysis of cellobiose (Atkins, Hooper & Isaac, 1973), based on steric hindrance, shows that less than 1% of the possible conformations from rotations of the glycosidic bonds (Pincus et al., 1976) are allowed when non-bonded interactions (Anfinsen & Scheraga, 1975) are considered.

flexibility. Not only are there theoretical arguments indicating that the conformations of nucleic acid polymers are able to rotate easily about the phosphodiester bond (Levitt, 1978) but experimentally it has been established that puckering of the sugar ring in the backbone distorts the shape of nucleic acids even in the doubly helical form. Condensation of random sequence RNA molecules to form catalytic units appears to be extremely unlikely. Specific base sequences would need to be favoured before three-dimensional, time stable structures would be formed from polynucleotides. In the absence of other catalytic molecules, nucleic acids are unlikely to form the basis of a self-replicating system. Even if simple

PREBIOTIC

EVOLUTION

25

polynucleotides such as poly A were produced prebiotically, base pairing would not lead to a self-ordering system. Indeed, it might be expected that extensive base pairing would simply neutralize the informational potential of the molecule. Obviously the initial polynucleotides would have been polydisperse both with respect to sequence and size, and without sufficient conformational integrity would be unlikely to generate catalytically active molecules. The molecular properties of nucleic acid suggests that these polymers may have arisen to create a more efficient molecular replica of a more rigid, less compact arrangement of molecular information, as could be expected in a polysaccharide template. The models of prebiotic evolution based on the spontaneous synthesis of deoxynucleotides, RNA or catalytic protenoids (Eigen et al., 1981) ignore the conformational properties of these molecules. Although tRNA molecules of today’s biosphere have complex secondary structure, the evidence that primordial RNA could have been synthesized abiotically or that the molecules could have had any semblance of a stable tertiary structure is not strong. How RNA, no matter what sequence or conformation could catalyze self-replication (Eigen et al., 1981), is still not clear. It is necessary to remind ourselves of the properties likely to allow abiotic “self-replication”. In an attempt to understand the relevance of the molecular interactions which define a living cell and which would be required to generate a self-ordering system, it is first necessary to consider briefly the properties of an organized system (Atlan, 1974). A simple formulation will suffice to illustrate some of the features the present model attempts to address. Atlan’s definition of organization: H(t)=Ho+t,+&

where H(t) is a measure of the organization at time t, Ho is the initial information, t,,, is a measure of the reliability and R0 is a measure of the initial structural redundancy; provides a conceptual framework which allows some of the important interactions in a living system to be visualized (Atlan, 1974). Our notion of a living system corresponds to this definition in that it requires a relationship between stored information (Ho; e.g. DNA) and the time-dependent synthesis (t,,,) and spatial distribution of ordered molecules (&; e.g. proteins or polysaccharides). Fundamental to our concept of life is that the organization content of the system is dependent on time. A change in the organization (i.e. evolution) is inevitable because the reliability of the molecular interactions is finite and dependent on exogenous influences. Atlan’s discussion on the requirements for the formation of a self-ordering system (Atlan, 1974) points to a possible route for

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prebiotic evolution based on a polymer with a high degree of structural redundancy. A consideration of the available building blocks, the conformational properties of their polymers and chemical reactivity appears to favor polysaccharides (Sharon & Lis, 1981). In the Beginning Before attempting to build a self-ordering macromolecular (polysaccharide) cycle, it is useful to consider properties and abundance of the first “organic” molecules. Many organic molecules have been detected in interstellar space (Mann & Williams, 1980), for example formaldehyde (Snyder et al., 1969), acetaldehyde (Ball, J. A., Gottlieb, C. A., Lilloy, A. E. & Radford, H. E., ht. Astron Union Circular No. 2350) and water (Cheung et al., 1969). These molecules are all chemically reactive and capable of forming both ordered crystalline arrays (e.g. ice) and organic polymers. The electronic structures are displayed in Fig. 2 to emphasize that the atoms and electron pairs within even a simple molecule have defined three-dimensional orientations. Acetaldehyde can be arranged in many possible conformations by rotation around the C-C single bond (Fig. 2) but intramolecular interactions favor one of these conformations. It is the simple extension of this principle which dominates the final conformations of even the most complex proteins or nucleic acids. A primative earth was a favorable environment for molecular-genesis. Many beautiful experiments have been performed to stimulate such conditions (Eck et al., 1966; Miller, 1953) and most have yielded interesting mixtures of organic molecules. A single example hardly suffices, but the experiment of Eck in 1966 where a mixture of CO2 and CH3 (1: 6) was heated at 500” K illustrates the enormous range of polyatomic organic molecules which can be synthesized even under simple conditions (Eck et al., 1966). Formaldehyde and acetaldehyde were detected at significant concentrations and even molecules such as cyclohexane and octanoic acid were present. Again the conformational properties of these molecules illustrate the forces which presumably controlled the evolution towards biomolecules. The space occupied by cyclohexane is dominated by the tetrahedral distribution of bonded atoms and the interactions between atoms across the six-membered ring. The tetrahedral arrangement of bonds around the carbon atom allows only two possible atomic arrangements for cyclohexane: a “boat” or a “chair”, but one of these conformations (the “chair”) allows the atoms to pack without interfering with each other. Octanoic acid is able to order space in a different way: the two ends of molecules such as this interact differently with water. These molecules tend

PREBIOTIC

27

EVOLUTION

Formaldehyde

Water

Acefaldehyde

FIG. 2. The shapes of polymers and polyatomic crystalline arrays emanate from the intrinsic spatial and conformational characteristics of their simplest constituents. When considering the structures and properties of nucleic acids and proteins, it is easy to become immersed in their complexity. However, it is the spatial distributions of electrons and nuclei in local regions of the molecules which determine the conformational and informational properties of the macromolecules. Formaldehyde is a reactive molecule capable of interacting with water to form hydrogen-bonds with preferential orientations. Water molecules interact in a nondirectional sense with methyl groups directing particular conformations and even long-range condensation (Anfinsen & Scheraga, 1975). The structure of the water lattice is also an important environmental feature to be considered when attempting to understand the attainment of native macromolecular conformations. Acetaldehyde has an internal degree of freedom to rearrange the position of its atoms by rotation around the C-C bond. Whilst a range of conformers between A and B is possible, the electronic interactions across the C-C bond favor the eclipsed arrangement (A). It is these same basic forces which determine the conformation of amino acid, nucleoside and saccharide units as well as the interactions between polymers. Molecular evolution must have been determined not only by chemical reactivity, but also to a great extent by the conformational determinants of the more abundant molecules.

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to form micelles and vesicles and, as a consequence, create different boundaries (or niches) within a water solution. It is interesting to note that fatty acids from Ca and Cs and even palmitic acid were detected in an analysis of samples from the Murchison meteorite (Krenvold, Lawless & Pormamperuma, 1971) and were almost certainly part of the early abiotic molecular repertoire. Sugars and Polysaccharides Considerable attention has been given to the formation of amino acids (Miller, 1953; Eck et al., 1966) and nucleosides (Lohrmann, 1976; Sherwood & Oro, 1977) under abiotic conditions. However, the most ubiquitous of biological molecules formed in these experiments (formaldehyde and acetaldehyde) are the precursors of monosaccharides. Formaldehyde and acetaldehyde have been subjected to a variety of ionizing radiations and base hydrolysis and in every case both pentoses and hexoses have been formed. As early as 1861, Butlerow was able to synthesize racemic mixtures of sugars such as glucose from formaldehyde (Butlerow, 1861). Evidence has been presented by Dhar and Ram (Dhar & Ram, 1932) that formaldehyde could form high in our atmosphere and reach concentrations as high as 1 mg/ml. Simulating the temperatures and surface conditions of hot mud springs has yielded racemic mixtures of riboses and deoxyriboses from formaldehyde solutions (Gabel & Ponnamperuma, 1967). Evolution towards a biological system necessitates the formation of either informational or structural polymers. It has been almost impossible to generate polynucleotides from simple precursors under abiotic conditions (Armstrong et al., 1977) and even the conditions for generating protenoids are rather vigorous (Temussi, 1976; Fox, 1976). However the formation of polysaccharides from simple sugars can be accomplished quite easily. Barker (1959) y-irradiated an aqueous solution of glucose, hydroxy acids and amino acids under anaerobic conditions and formed polysaccharides (Barker et al., 1959). These polymerizations do not always appear to generate polydisperse products. By heating glucose at 50-60°C in dimethylformamide (or in water at 175”C, Mora & Wood, 1958), it was possible to synthesize a poly 1, 6-glycoside with an average molecular weight of almost 30 000 daltons (Schramm, Griitsch & Pollman, 1962). Why consider polysaccharides as the primordial spark for biological evolution? There are four properties of polysaccharides which suggest a role for them in prebiotic evolution: (a) they are conformationally restricted (i.e.

PREBIOTIC

EVOLUTION

29

simple polysaccharides form helical structures in three dimensions), (b) the conformations appear to contain considerable redundancy (i.e. changes in the nature of the sugar units do not necessarily destroy the conformational preferences of the polysaccharide backbone), (c) the repetitive formation of a given polysaccharide structural domain could occur reliably in the absence of a unique catalyst (this is a result of the conformational properties of the glycosidic linkage, see later) and (d) oligosaccharides can act as powerful catalysts (van Hooidonk & Breebaart-Hansen, 1970; Tutt & Schwartz, 1971). It is useful to consider the properties of polysaccharides with reference to Atlan’s definition of organization given earlier: the concept of initial information (Ho) can be equated with the conformational restriction; the redundancy (&) can be associated with both the ability of polysaccharides to form similar three-dimensional structures (despite side chain substitution of the monosaccharides) and the helices which form repetitive structural domains; and finally, the reliability (t,,,) of formation of these helices is directly related to the conformational properties of the short-range atomic interactions across the glycosidic bond (Pincus, Burgess & Scheraga, 1976). It is important to compare the conformational properties of polysaccharides, polyamino acids and polynucleotides when assessing their roles in the initiation of a self-ordering cycle. Polypeptides occurring in abiotic times were presumably composed of different amino acids in random sequence array. No experiments have demonstrated that a polymer with three-dimensional integrity can arise from such polyamino acids, although some unusual amino acids (e.g. a aminoisobutyric acid) have highly directional conformational information (Burgess & Leach, 1973) and may have directed the synthesis of small peptides with conformational integrity. These amino acids have not survived as units in our genetic system so their role in initial protenoids may have been obscured. Some organisms still use these amino acids to construct biologically active peptides independently of direct nucleic acid involvement (Payne, Jakes & Hartley, 1970). When it is considered that glycine was probably the most abundant amino acid, the conformational flexibility (Fig. 1) of these polymers would have been too great to allow the repeated production of a three-dimensional structure able to catalyze its own formation. The difference in the conformational flexibility on either side of the peptide bond and the glycosidic bond can be compared using the respective 4, $ energy diagrams (Fig. 1). This limited range of conformations for the glycosidic bond inevitably leads to the formation of highly ordered helical polymers such as the starches, chitins or celluloses. Although biological polysaccharides often consist of a single enantiomer, the initial polymers would have been formed

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from racemic mixtures of L- and D-sugars. The conformations generated from racemic mixtures would still be expected to form rather rigid conformations in which the repeated production of catalytic sites could occur. At later stages in evolution, the repertoire of conformations available to proteins would have allowed the generation of more powerful catalysts to supercede the catalytic function of the original carbohydrates.

Catalysis Conceptually, abiotic experiments have been directed towards the generation of catalysts which would increase the reliability of a given molecular cycle (Fox & Dose, 1972). The discussion above indicates that some polysaccharides may develop three-dimensional order in the absence of catalysts of high specificity. However oligosaccharides (especially oligosaccharides doped with dyes, Yushina, 1957) can also be catalysts. A most exciting and beautiful example of saccharide catalysis was discovered using several cycloamyloses (van Hooidonk & Breebaart-Hansen, 1970; Tutt & Schwartz, 1971). The cycloamyloses demonstrate substrate specificity (van Hooidonk & Breebaart-Hansen, 1970) and their rate of catalysis depends intimately on the three-dimensional shape of the cycloamylose. A seven membered oligosaccharide ring is able to increase the rate of hydrolysis of m-tert-butyl phenylacetate 100 OOO-fold compared to the rate of hydrolysis by the hydroxyl ion alone. Cyclohexaamylose on the other hand only increases the rate of hydrolysis 5000-fold. Enhancement of a reaction rate to this extent is comparable to enzymic catalysis by (Y chymotrypsin where the enhancement factor is lo6 (Hennrich & Cramer, 1965). Cycloheptaamylose catalyzed hydrolysis can be increased a further three-fold when an imidazole group is attached to the sugar ring (Cramer & Mackensen, 1966). It can be assumed that the fidelity of the initial replicative events was rather low. However, the synthesis of a given cyclical or helical polysaccharide conformation would be expected to occur continuously and the range of catalytic activities could have been quite diverse. Obviously other catalysts may have arisen simultaneously, thus initiating a variety of hypercyclic systems. Since this discussion is directed towards events “beyond” measurable time, these possibilities will not be expanded upon, except to say that the other catalysts may have been organometallic molecules, e.g. hematoporphyrins (Szukta, Hazel & McNab, 1959) or organic complexes of iron and sulphur such as the ones which occur in some bacterial proteins (Lane et al., 1975; Adam, Sieker & Jensen, 1973; Holm, 1975), both of which form easily under abiotic conditions.

PREBIOTIC

Phases-A

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EVOLUTION

Primitive

Cell

Two more fundamental properties of living systems are related to their phase separation from the surrounding milieu. Firstly, biomolecular reactions occur within a protected environment (a cell), where the molecules entering and leaving are biased by the nature of the phase barrier (membrane or cell wall). Secondly, the system grows with time to a critical level of organization and then divides almost equally to permit the proliferation of this spatial organization. Oscillatory growth and replication was presumably a very early part of the prebiotic process. The most attractive molecules for the initiation of phase separations and membrane formation are glycolipids. Both the lipid and carbohydrate components of glycolipids can be assumed to be present in the abiotic era, and glycolipids are a ubiquitous feature of all biological systems today. Glycolipids isolated from ancient organisms such as the blue green algae form micelles with remarkable diffusion and stability properties (Lambein & Walk, 1973). These glycolipids are able to form laminated micelles, and, in combination with the polysaccharide complex isolated from the same organism (Lambein & Wolk, 1973; CardemiI & Walk, 1979), allow the heterocysts to exclude O2 and to withstand boiling water. The lamella vesicles formed by lipids and glycolipids of the type found in the blue algae have even been shown to selectively accumulate particular ions and amino acids (Armstrong et aE., 1977). A scenario for the formation of a possible initial self-ordering system is illustrated in Fig. 3. Initially sugars and fatty acids accumulated and combined to form both polysaccharides and glycolipids. The glycolipids formed micelles and vesicles which sequestered or promoted the internal synthesis of polysaccharides + these vesicles favored the concentration of monosaccharides which reacted to form polysaccharides + some of the polysaccharides were conformationally ordered catalyzing an accumulation of similar polymers + as the concentration of the hydrophilic polysaccharides increased, the vesicles destabilized and regularly broke down into smaller units. Such a system would be expected to be structured, oscillatory and reasonably stable with respect to time. However, given that a polysaccharide will repeatedly form helical arrays or highly similar repetitive molecular environments, which could be expected to provide a catalytic environment (especially when the polysaccharide was “seeded” with side chains such as nucleic acid bases), it is reasonable to propose that one such domain catalyzed the formation of similar glycosidic linkages, i.e. the formation of self. Even if this catalytic activity was weak, such a system would then be expected to replicate with an enormous

FIG. 3. The initiation of a self-ordering molecular system can be depicted by the growth and division of a glycolipid-polysaccharide vesicle. Initiation of the cycle takes place through the entrapment of conformationally similar polysaccharide molecules (A, B, C, D, E). Within the vesicular environment, the initial polysaccharide may simply have stabilized the vesicle. The abundance of such vesicles would then be expected to increase with time and some vesicles would be expected to exhibit selective permeability to monosaccharides (0,O) or amino acids (A, A). A particular polysaccharide may have exhibited weak catalytic activity (A, B*, C, D, E) favoring the polymerization of similar polymers. Such catalysis would increase the abundance of such vesicles, given an excess of glycolipid (e) for vesicle formation. When the catalysis was sufficient to build up a significant intramicellar polymer concentration, the vesicle may be expected to divide, i.e. become a self-reproducing unit equivalent to a primordial cell.

*A

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advantage. If the initial catalysis involved the side chains on such a polysaccharide, a dependency of this activity on particular bases could have risen. Presumably the bases would have been scattered on the surface of the polysaccharide, but accumulation of bases at the helical turns or other structural pockets may have provided a unique informational template from which the self-replicating genome could have developed. It is tempting to pause and leave the mode of evolution of today’s genetic code from this point to the reader’s speculation-for there are a myriad of ways to proceed and it is a little more than adventurous to commit oneself to a particular scenario. But there are some appealing speculations which can relate the structure, arrangement and expression of the information in our current genome to this initial model for prebiotic evolution. Although self -ordering and cycling, the initial polysaccharide-glycolipid system is structurally rigid. The informational properties would be dependent on the generation of restricted conformations, the formation of which would result from the conformational properties of the glycosidic bonds. Although side chains in the form of bases (such links are still present in many antibiotics, Suhadolnik, 1970) may have increased the reliability of template “replication”, the efficiency of reproduction would be expected to be low. When the polysaccharide template developed a catalytic activity, which aided the polymerization of amino acids, the start of a new, and more powerful self-ordering cycle can be envisaged. The first polyamino acids may have conferred little or no selective advantage to the polysaccharide glycolipid system, but by chance a protein which stabilized the micelle or facilitated the synthesis of the polysaccharide template may have been produced. Such polyamino acids need not have had strict sequence integrity as we know it, but may have acted by providing charge shielding (e.g. histone-like) or regions of lower dielectric constant to facilitate catalysis. However, particular configurations of the polysaccharide and its side chains (including nucleotides juxtaposed by virtue of being on adjacent helical turns) might have been expected to favor particular amino acids. The first protein polysaccharide hypercycle could thus have arisen spontaneously as a result of polysaccharide catalysis (Eigen, 1971). Despite the distance in time between the initial self-ordering system and the intricate cellular networks of today, it is interesting to note that polysaccharides still play a vital role in the structure of the eukaryotic nuclei. It has been shown that all nuclei contain polysaccharides (Mohberg & Rusch, 1971) and nucleopolysaccharides (e.g. poly ADP-ribose, Rechsteiner, Hillyard & Olivera, 1976), but much remains to be known about the structural or functional role of these nuclear polysaccharides (excepting DNA)

34 which constitute by weight, (Mohberg & Rusch, 1971).

A.

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almost

as much nuclear polymer

as DNA

Continuity

It will be dithcult to trace a unique (if one existed) pathway between the present day genome and the abiotic molecular state. However, one principle which would appear to be valid is that there has been a continuous transfer of structural redundancy and reliability (as defined by molecular conformations) into informational storage from which the structural requirements are subsequently generated. This corresponds thermodynamically to a continuous increase in the potential entropy of a system (i.e. as the informational content of the system increases, so does the entropy, Wilson, 1968). Indeed, it may be rationally argued that it is the thermodynamic drive to increase the informational content of a molecular system which answers the question: “Why evolution?“. Thus the early polysaccharides may have been replaced by a flexible polynucleotide (i.e. nucleic acid) capable of mimicking the catalytic niches for synthesizing polyamino acids more efficiently. Since the initial information must have been dispersed over the surface of the polysaccharide, one would expect the polymer which replicated this information to be dispersed as well (Fig. 4) (i.e. most of the relevant information would be separated by bases which served only to link the informational niches). Our present understanding of the eukaryotic genome shows it to be dispersed in this sense-i.e. much of the coding region is interrupted by untranslated sequences of nucleotides. Of course, the eukaryotic evolution may have made use of these intron linkages to allow increased flexibility in the expression of usefully encoded domains to produce new proteins. Presumably viruses and bacteria have evolved by eliminating the linkage sequences, thus increasing the efficiency of information storage. The evolution of intermediary steps in the development of information transfer from a nucleic acid template to structural and functional proteins presumably increased the efficiency and reliability of the process. However, the pathway of biomolecular evolution must also have been influenced by two further features of cells as we know them today: firstly, the transfer of the protein synthesizing machinery from the site of transcription to a distant site within the cell; and secondly, the inevitable oscillatory mode which re-organizes the system, regularly leading to replication (Winfree, 1978). In the first case, both mRNA’s and tRNA’s may have evolved structural sequences which would direct the diffusion of these molecules away from the nucleus into the cytoplasm.

PREBIOTIC

>< Catalytic

(b)

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EVOLUTION

domains

‘Protein’

FIG. 4. An initial polymeric catalyst was unlikely to be homogeneous with respect to composition, or conformation. However, given the propensity of complex polysaccharides to form similar conformations (including helices), particular niches might have been expected to be repeated at different sites on the surface of the molecule. Such sites may have had chemically “reactive” environments such as occurred in the cycloamyloses (Hennrich & Cramer, 1965) or catalytic environments created by the absorption (Yushina, 1957) of other molecules such as the porphyrins (Szukta et al., 1959). Whilst these sites could be expected to be repeated, the relatively infrequent generation of a specific catalytic activity (directed by conformational determinants) may be expected to reproduce with low efficiency. However as the abundance of catalytic activity increased, more efficient replicas of the catalytic sites would be expected to emerge. Such replicates can be imagined as the prototypes of “informational” polymers with introns and exons.

Indeed it is interesting to speculate that introns may have a functional role to direct the folding of the intitial mRNA so that it remains in the nucleus until other cellular processes have been completed and the cell is ready to use the mRNA product. This would allow both spatial and temporal organization of molecular events (Winfree, 1978) so essential for the maintenance of cellular integrity (cf. proteolytic cleavage during viral morphogenesis). Studies on the rate of diffusion of macromolecules in gel networks (Cumming, Handley & Preston, 1979; Laurent, Preston &

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Sundeliif, 1979) suggest that the hydrodynamic properties of macromolecules may be important for controlling the distribution of these molecules within the various subcellular compartments, Thus, evolutionary changes in nucleotide sequences may not necessarily be directly related to the structure of a protein, but for the determination of the spatial distribution of a mRNA or its precursor and thus allow control over the time within the cell cycle at which such a message is translated. Obviously it is only possible to address a miniscule proportion of the questions which arise during a consideration of prebiotic evolution. Many aspects remain neglected, for example considerable experimental effort has been directed towards an explanation of the reason for the asymmetric chirality of amino acids in proteins (Edwards, Cooper & Dougherty, 1980). In spite of some extraordinary intellectual slides (Asimov, 1976) the case for overall achirality in our molecular “universe” is convincingly presented by others (Frank, 1953; Wald, 1957; Swan, 1979). One final consideration of the events later in evolutionary development also suggests further possibilities for the properties of our present genome. The existence of these organometallic “catalysts” and their incorporation into the active sites of proteins is surely a reflection on the versatility of the information template in living systems. Given that complexes such as Fe-!& (Lane et al., 1975; Holm, 1975) existed before their existence in a protein (Adman et al., 1973), it is intriguing to consider how such proteins could have evolved. One possible mechanism is that the genome has the capability of synthesizing a protein which is able to entrap the Fe-!% molecule, Given the existence of the immune system which can recognize a vast range of antigens, it is interesting to consider the possibility that there are (or were) other protein recognition systems. Such systems would be expected to allow for a repertoire of molecules to be synthesized in response to the presence of an inducing molecule. Such a recognition system might be expected to be based on a set of conformational domains, capable of achieving protein globularity, but which allowed flexibility in ligand complexing. Whilst the initial successful products may have been active as protein-co-ordinate complexes, further evolution might be expected to allow the production of a protein which also forms part of the active organometallic complex. If such a recognition repertoire exists, even the present day genome might be expected to contain several copies of a particular domain of a given protein, e.g. the active gene copy, the recognition repertoire copy and even vestigial copies of initial, partially successful attempts to produce proteins in response to an inductive stimulus. These latter copies, if not efficiently eliminated, could be detected as the pseudo genes reported for molecules such as globin (Haynes et al., 1980).

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