New prospects for deducing the evolutionary history of metabolic pathways in prokaryotes: Aromatic biosynthesis as a case-in-point

Metabolic pathways of prokaryotes are more biochemically diverse than is generally recognized. Distinctive biochemical features are shared by phylogenetic clusters. The hierarchical levels of characterstate clustering depends upon evolutionary events which fortuitously became fixed in the genome of a common ancestor. Prokaryotes can now be ordered on a phylogenetic tree. This allows the evolutionary steps that underlie the construction and regulation of appropriately complex biochemical pathways to be traced in an evolutionary progression of prokaryote types that house these pathways. Essentially the approach is to deduce ancestral character states at ever deeper phylogenetic levels, utilizing logical principles of maximum parsimony. The current perspective on the evolution of the biochemical pathway for biosynthesis of aromatic amino acids is developed as a case-in-point model for analyses that should be feasible with many major metabolic systems. Phenylalanine biosynthesis probably arose prior to the addition of branches leading to tyrosine and tryptophan. An evolutionary scenario is developed that begins with non-enzymatic reactions which may have operated in primitive systems, followed by the evolution of an enzymatic system that pre-dated the divergence of major lineages of modern eubacteria (Gram-positive bacteria, Gram-negative purple bacteria, and cyanobacteria).Florida Agricultural Experiment Station, Journal Series No. 8251.     

The tragedy of the commons in microbial populations: insights from theoretical, comparative and experimental studies

First principles of thermodynamics imply that metabolic pathways are faced with a trade-off between the rate and yield of ATP production. Simple evolutionary models argue that this trade-off generates a fundamental social conflict in microbial populations: average fitness in a population is highest if all individuals exploit common resources efficiently, but individual reproductive rate is maximized by consuming common resources at the highest possible rate, a scenario known as the tragedy of the commons. In this paper, I review studies that have addressed two key questions: What is the evidence that the rate-yield trade-off is an evolutionary constraint on metabolic pathways? And, if so, what determines evolutionary outcome of the conflicts generated by this trade-off? Comparative studies and microbial experiments provide evidence that the rate-yield trade-off is an evolutionary constraint that is driven by thermodynamic constraints that are common to all metabolic pathways and pathway-specific constraints that reflect the evolutionary history of populations. Microbial selection experiments show that the evolutionary consequences of this trade-off depend on both kin selection and biochemical constraints. In well-mixed populations with low relatedness, genotypes with rapid and efficient metabolism can coexist as a result of negative frequency-dependent selection generated by density-dependent biochemical costs of rapid metabolism. Kin selection can promote the maintenance of efficient metabolism in structured populations with high relatedness by ensuring that genotypes with efficient metabolic pathways gain an indirect fitness benefit from their competitive restraint. I conclude by suggesting avenues for future research and by discussing the broader implications of this work for microbial social evolution.     

The tragedy of the commons in microbial populations: insights from theoretical, comparative and experimental studies

First principles of thermodynamics imply that metabolic pathways are faced with a trade-off between the rate and yield of ATP production. Simple evolutionary models argue that this trade-off generates a fundamental social conflict in microbial populations: average fitness in a population is highest if all individuals exploit common resources efficiently, but individual reproductive rate is maximized by consuming common resources at the highest possible rate, a scenario known as the tragedy of the commons. In this paper, I review studies that have addressed two key questions: What is the evidence that the rate-yield trade-off is an evolutionary constraint on metabolic pathways? And, if so, what determines evolutionary outcome of the conflicts generated by this trade-off? Comparative studies and microbial experiments provide evidence that the rate-yield trade-off is an evolutionary constraint that is driven by thermodynamic constraints that are common to all metabolic pathways and pathway-specific constraints that reflect the evolutionary history of populations. Microbial selection experiments show that the evolutionary consequences of this trade-off depend on both kin selection and biochemical constraints. In well-mixed populations with low relatedness, genotypes with rapid and efficient metabolism can coexist as a result of negative frequency-dependent selection generated by density-dependent biochemical costs of rapid metabolism. Kin selection can promote the maintenance of efficient metabolism in structured populations with high relatedness by ensuring that genotypes with efficient metabolic pathways gain an indirect fitness benefit from their competitive restraint. I conclude by suggesting avenues for future research and by discussing the broader implications of this work for microbial social evolution.     

The possible role of methylglyoxal metabolism in cancer

Tumours reprogram their metabolism to acquire an evolutionary advantage over normal cells. However, not all such metabolic pathways support energy production. An example of these metabolic pathways is the Methylglyoxal (MG) one. This pathway helps maintain the redox state, and it might act as a phosphate sensor that monitors the intracellular phosphate levels. In this work, we discuss the biochemical step of the MG pathway and interrelate it with cancer.     

Ordering events of biochemical evolution

Metabolic pathways exhibit structures resulting from an evolutionary process. Pathways have been inherited through time with modification, from the earliest periods of life. It is possible to compare the structure of pathways as done in comparative anatomy, i.e. for inferring ancestral pathways or parts of it (ancestral enzymatic functions), using standard phylogenetic reconstruction. Thus a phylogenetic tree of pathways provides a relative ordering of the rise of enzymatic functions. It even becomes possible to order the birth of each complete pathway in time. This particular "DNA-free" conceptual approach to evolutionary biochemistry is reviewed, gathering all the justifications given for it. Then, the method of assigning a given pathway to a time span of biochemical development is revisited. The previous method used an implicit "clock" of metabolic development that is difficult to justify. We develop a new clock-free approach, using functional biochemical arguments. Results of the two methods are not significantly different; our method is just more precise. This suggests that the clock assumed in the first method does not provoke any important artefact in describing the development of biochemical evolution. It is just unnecessary to postulate it. As a result, most of the amino acid metabolic pathways develop forwards, confirming former models of amino acid catabolism evolution, but not those for amino acid anabolism. The order of appearance of sectors of universal cellular metabolism is: (1) amino acid catabolism, (2) amino acid anabolism and closure of the urea cycle, (3) glycolysis and glycogenesis, (4) closure of the pentose-phosphate cycle, (5) closure of the Krebs cycle and fatty acids metabolism, (6) closure of the Calvin cycle.     

Patterns of evolutionary rate variation among genes of the anthocyanin biosynthetic pathway

The anthocyanin biosynthetic pathway is responsible for the production of anthocyanin pigments in plant tissues and shares a number of enzymes with other biochemical pathways. The six core structural genes of this pathway have been cloned and characterized in two taxonomically diverse plant species (maize and snapdragon). We have recently cloned these genes for a third species, the common morning glory, Ipomoea purpurea. This additional information provides an opportunity to examine patterns of evolution among genes within a single biochemical pathway. We report here that upstream genes in the anthocyanin pathway have evolved substantially more slowly than downstream genes and suggest that this difference in evolutionary rates may be explained by upstream genes being more constrained because they participate in several different biochemical pathways. In addition, regulatory genes associated with the anthocyanin pathway tend to evolve more rapidly than the structural genes they regulate, suggesting that adaptive evolution of flower color may be mediated more by regulatory than by structural genes. Finally, for individual anthocyanin genes, we found an absence of rate heterogeneity among three major angiosperm lineages. This rate constancy contrasts with an accelerated rate of evolution of three CHS-like genes in the Ipomoea lineage, indicating that these three genes have diverged without coordinated adjustment by other pathway genes.     

The alphaD-globin gene originated via duplication of an embryonic alpha-like globin gene in the ancestor of tetrapod vertebrates

Gene duplication is thought to play an important role in the co-option of existing protein functions to new physiological pathways. The globin superfamily of genes provides an excellent example of the kind of physiological versatility that can be attained through the functional and regulatory divergence of duplicated genes that encode different subunit polypeptides of the tetrameric hemoglobin protein. In contrast to prevailing views about the evolutionary history of the alpha-globin gene family, here we present phylogenetic evidence that the alpha(A)- and alpha(D)-globin genes are not the product of a single, tandem duplication of an ancestral globin gene with adult function in the common ancestor of extant birds, reptiles, and mammals. Instead, our analysis reveals that the alpha(D)-globin gene of amniote vertebrates arose via duplication of an embryonic alpha-like globin gene that predated the radiation of tetrapods. The important evolutionary implication is that the distinct biochemical properties of alpha(D)-hemoglobin (HbD) are not exclusively derived characters that can be attributed to a post-duplication process of neofunctionalization. Rather, many of the distinct biochemical properties of HbD are retained ancestral characters that reflect the fact that the alpha(D)-globin gene arose via duplication of a gene that had a larval/embryonic function. These insights into the evolutionary origin of HbD illustrate how adaptive modifications of physiological pathways may result from the retention and opportunistic co-option of ancestral protein functions.     

Metabolic systems cost-benefit analysis for interpreting network structure and regulation

MOTIVATION: Interpretation of bioinformatics data in terms of cellular function is a major challenge facing systems biology. This question is complicated by robust metabolic networks filled with structural features like parallel pathways and isozymes. Under conditions of nutrient sufficiency, metabolic networks are well known to be regulated for thermodynamic efficiency however; efficient biochemical pathways are anabolically expensive to construct. While parameters like thermodynamic efficiency have been extensively studied, a systems-based analysis of anabolic proteome synthesis 'costs' and the cellular function implications of these costs has not been reported. RESULTS: A cost-benefit analysis of an in silico Escherichia coli network revealed the relationship between metabolic pathway proteome synthesis requirements, DNA-coding sequence length, thermodynamic efficiency and substrate affinity. The results highlight basic metabolic network design principles. Pathway proteome synthesis requirements appear to have shaped biochemical network structure and regulation. Under conditions of nutrient scarcity and other general stresses, E. coli expresses pathways with relatively inexpensive proteome synthesis requirements instead of more efficient but also anabolically more expensive pathways. This evolutionary strategy provides a cellular function-based explanation for common network motifs like isozymes and parallel pathways and possibly explains 'overflow' metabolisms observed during nutrient scarcity. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.     

Evolutionary innovation and diversification of carotenoid‐based pigmentation in finches

The ornaments used by animals to mediate social interactions are diverse, and by reconstructing their evolutionary pathways we can gain new insights into the mechanisms underlying ornamental innovation and variability. Here, we examine variation in plumage carotenoids among the true finches (Aves: Fringillidae) using biochemical and comparative phylogenetic analyses to reconstruct the evolutionary history of carotenoid states and evaluate competing models of carotenoid evolution. Our comparative analyses reveal that the most likely ancestor of finches used dietary carotenoids as yellow plumage colorants, and that the ability to metabolically modify dietary carotenoids into more complex pigments arose secondarily once finches began to use modified carotenoids to create red plumage. Following the evolutionary “innovation” that enabled modified red carotenoid pigments to be deposited as plumage colorants, many finch species subsequently modified carotenoid biochemical pathways to create yellow plumage. However, no reversions to dietary carotenoids were observed. The finding that ornaments and their underlying mechanisms may be operating under different selection regimes—where ornamental trait colors undergo frequent reversions (e.g., between red and yellow plumage) while carotenoid metabolization mechanisms are more conserved—supports a growing empirical framework suggesting different evolutionary patterns for ornaments and the mechanistic innovations that facilitate their diversification.     

Dishevelled: at the crossroads of divergent intracellular signaling pathways.

During the development of multicellular organisms the formation of complex patterns relies on specific cell-cell signaling events. For tissues to become spatially organized and cells to become committed to specialized fates it is absolutely crucial for proper development that the underlying signaling systems receive and route information correctly. Recently, a wealth of genetic and biochemical experimental data has been collected about prevalent evolutionary conserved signaling families, such as the Wnts, Dpp/BMPs, and Hedgehogs, in flies, worms, and vertebrates. Paradoxically, members of a particular signaling family often have receptors with similar biochemical binding properties, though they activate different intracellular pathways in vivo and can be phenotypically distinguished. How are their specific biological responses then generated? With respect to signaling specificity in Wnt pathways, Dishevelled is an intriguing protein; in Drosophila melanogaster it is required in two distinct signaling pathways, that share Frizzled receptors of similar structure, but have distinct intracellular signaling routes. Recent results suggest that Dishevelled is a multifunctional protein at the crossroads of divergent Wnt/Fz pathways. Dishevelled appears to be a key factor in Wnt signaling to read' signals coming from the plasma membrane and route them into the correct intracellular pathways.     

libSRES: a C library for stochastic ranking evolution strategy for parameter estimation

SUMMARY: Estimation of kinetic parameters in a biochemical pathway or network represents a common problem in systems studies of biological processes. We have implemented a C library, named libSRES, to facilitate a fast implementation of computer software for study of non-linear biochemical pathways. This library implements a (mu, lambda)-ES evolutionary optimization algorithm that uses stochastic ranking as the constraint handling technique. Considering the amount of computing time it might require to solve a parameter-estimation problem, an MPI version of libSRES is provided for parallel implementation, as well as a simple user interface. libSRES is freely available and could be used directly in any C program as a library function. We have extensively tested the performance of libSRES on various pathway parameter-estimation problems and found its performance to be satisfactory. AVAILABILITY: The source code (in C) is free for academic users at http://csbl.bmb.uga.edu/~jix/science/libSRES/     

The role of lateral gene transfer in the evolution of isoprenoid biosynthesis pathways

Lateral gene transfer (LGT) is a major force in microbial genome evolution. Here, we present an overview of lateral transfers affecting genes involved in isopentenyl diphosphate (IPP) synthesis. Two alternative metabolic pathways can synthesize this universal precursor of isoprenoids, the 1-deoxy-D-xylulose 5-phosphate (DOXP) pathway and the mevalonate (MVA) pathway. We have surveyed recent genomic data and the biochemical literature to determine the distribution of the genes composing these pathways within the bacterial domain. The scattered distribution observed is incompatible with a simple scheme of vertical transmission. LGT (among and between bacteria, archaea and eukaryotes) more parsimoniously explains many features of this pattern. This alternative scenario is supported by phylogenetic analyses, which unambiguously confirm several cases of lateral transfer. Available biochemical data allow the formulation of hypotheses about selective pressures favouring transfer. The phylogenetic diversity of the organisms involved and the range of possible causes and effects of these transfer events make the IPP biosynthetic pathways an ideal system for studying the evolutionary role of LGT.     

Two unique cyanobacteria lead to a traceable approach of the first appearance of oxygenic photosynthesis

The evolutionary route from anoxygenic photosynthetic bacteria to oxygenic cyanobacteria is discontinuous in terms of photochemical/photophysical reaction systems. It is difficult to describe this transition process simply because there are no recognized intermediary organisms between the two bacterial groups. Gloeobacter violaceus PCC 7421 might be a model organism that is suitable for analysis because it still possesses primordial characteristics such as the absence of thylakoid membranes. Whole genome analysis and biochemical and biophysical surveys of G. violaceus have favored the hypothesis that it is an intermediary organism. On the other hand, species differentiation is an evolutionary process that could be driven by changes in a small number of genes, and this process might give fair information more in details by monitoring of those genes. Comparative studies of genes, including those in Acaryochloris marina MBIC 11017, have provided information relevant to species differentiation; in particular, the acquisition of a new pigment, chlorophyll d, and changes in amino acid sequences have been informative. Here, based on experimental evidence from these two species, we discuss some of the evolutionary pathways for the appearance and differentiation of cyanobacteria.     

Use of Game-Theoretical Methods in Biochemistry and Biophysics

Evolutionary game theory can be considered as an extension of the theory of evolutionary optimisation in that two or more organisms (or more generally, units of replication) tend to optimise their properties in an interdependent way. Thus, the outcome of the strategy adopted by one species (e.g., as a result of mutation and selection) depends on the strategy adopted by the other species. In this review, the use of evolutionary game theory for analysing biochemical and biophysical systems is discussed. The presentation is illustrated by a number of instructive examples such as the competition between microorganisms using different metabolic pathways for adenosine triphosphate production, the secretion of extracellular enzymes, the growth of trees and photosynthesis. These examples show that, due to conflicts of interest, the global optimum (in the sense of being the best solution for the whole system) is not always obtained. For example, some yeast species use metabolic pathways that waste nutrients, and in a dense tree canopy, trees grow taller than would be optimal for biomass productivity. From the viewpoint of game theory, the examples considered can be described by the Prisoner’s Dilemma, snowdrift game, Tragedy of the Commons and rock–scissors–paper game.     

Evolutionary origins of metabolic compartmentalization in eukaryotes

Many genes in eukaryotes are acquisitions from the free-living antecedents of chloroplasts and mitochondria. But there is no evolutionary ‘homing device’ that automatically directs the protein product of a transferred gene back to the organelle of its provenance. Instead, the products of genes acquired from endosymbionts can explore all targeting possibilities within the cell. They often replace pre-existing host genes, or even whole pathways. But the transfer of an enzymatic pathway from one compartment to another poses severe problems: over evolutionary time, the enzymes of the pathway acquire their targeting signals for the new compartment individually, not in unison. Until the whole pathway is established in the new compartment, newly routed individual enzymes are useless, and their genes will be lost through mutation. Here it is suggested that pathways attain novel compartmentation variants via a ‘minor mistargeting’ mechanism. If protein targeting in eukaryotic cells possesses enough imperfection such that small amounts of entire pathways continuously enter novel compartments, selectable units of biochemical function would exist in new compartments, and the genes could become selected. Dual-targeting of proteins is indeed very common within eukaryotic cells, suggesting that targeting variation required for this minor mistargeting mechanism to operate exists in nature.     

Pathway Length and Evolutionary Constraint in Amino Acid Biosynthesis

The evolutionary properties of a metabolic network may be determined by the topology of the network. One attribute of pathways that make up the network is the number of enzymatic steps between initial substrates and final products. To determine the effect of pathway length on evolutionary lability of pathway structure, we examined amino acid biosynthetic pathways across 48 sequenced organisms. We demonstrate that longer pathways exhibit lower rates of change in pathway structure than shorter pathways. This finding suggests that increasing complexity may increase constraint on evolutionary change. (Matthew T. Rutter and Rebecca A. Zufall) Both authors contributed equally to this work.     

Protein transport in Archaea: Sec and twin arginine translocation pathways

The transport of proteins into and across hydrophobic membranes is an essential cellular process. The majority of proteins that are translocated in an unfolded conformation traverse the membrane by way of the universally conserved Sec pathway, whereas the twin arginine translocation pathway is responsible for the transport of folded proteins across the membrane. Structural, biochemical and genetic analyses of these processes in Archaea have revealed unique archaeal features, and have also provided a better understanding of these pathways in organisms of all domains. Further study of these pathways in Archaea might elucidate fundamental principles involved in each type of transport and could help determine their relative costs and benefits as well as evolutionary adaptations in protein secretion strategies.     

Fundamental Escherichia coli biochemical pathways for biomass and energy production: identification of reactions

Cells grow by oxidizing nutrients using a complex network of biochemical reactions. During this process new biological material is produced along with energy used for maintaining cellular organization. Because the metabolic network is highly branched, these tasks can be accomplished using a wide variety of unique reaction sequences. However, evolutionary pressures under carbon-limited growth conditions likely select organisms that utilize highly efficient pathways. Using elementary-mode analysis, we demonstrate that the metabolism of the bacterium Escherichia coli contains four unique pathways that most efficiently convert glucose and oxygen into new cells and maintenance energy under any level of oxygen limitation. Observed regulatory patterns and experimental findings suggest growing cells use these highly efficient pathways. It is predicted that five knockout mutations generate a strain that supports growth using only the most efficient reaction sequence. The analysis approach should be generally useful for predicting metabolic capabilities and efficient network designs based on only genomic information.     

Automated system for gene annotation and metabolic pathway reconstruction using general sequence databases

Despite the growing number of genomes published or currently being sequenced, there is a relative paucity of software for functional classification of newly discovered genes and their assignment to metabolic pathways. Available software for such analyses has a very steep learning curve and requires the installation, configuration, and maintenance of large amounts of complex infrastructure, including complementary software and databases. Many such tools are restricted to one or a few data sources and classification schemes. In this work, we report an automated system for gene annotation and metabolic pathway reconstruction (ASGARD), which was designed to be powerful and generalizable, yet simple for the biologist to install and run on centralized, commonly available computers. It avoids the requirement for complex resources such as relational databases and web servers, as well as the need for administrator access to the operating system. Our methodology contributes to a more rapid investigation of the potential biochemical capabilities of genes and genomes by the biological researcher, and is useful in biochemical as well as comparative and evolutionary studies of pathways and networks.