Yeast has evolved to minimize protein resource cost for synthesizing amino acids.

Proteins, as essential biomolecules, account for a large fraction of cell mass, and thus the synthesis of the complete set of proteins (i.e., the proteome) represents a substantial part of the cellular resource budget. Therefore, cells might be under selective pressures to optimize the resource costs for protein synthesis, particularly the biosynthesis of the 20 proteinogenic amino acids. Previous studies showed that less energetically costly amino acids are more abundant in the proteomes of bacteria that survive under energy-limited conditions, but the energy cost of synthesizing amino acids was reported to be weakly associated with the amino acid usage in Saccharomyces cerevisiae Here we present a modeling framework to estimate the protein cost of synthesizing each amino acid (i.e., the protein mass required for supporting one unit of amino acid biosynthetic flux) and the glucose cost (i.e., the glucose consumed per amino acid synthesized). We show that the logarithms of the relative abundances of amino acids in S. cerevisiae's proteome correlate well with the protein costs of synthesizing amino acids (Pearson's r = -0.89), which is better than that with the glucose costs (Pearson's r = -0.5). Therefore, we demonstrate that S. cerevisiae tends to minimize protein resource, rather than glucose or energy, for synthesizing amino acids.

Biosynthetic energy cost of potentially highly expressed proteins vary with niche in selected actinobacteria.

Amino acid and protein biosynthesis requires a number of high energy phosphate bonds and includes a dual energy cost for the synthesis of chemical intermediates during the fueling reactions and the conversion of precursor molecules to final products. One popular hypothesis is that the proteins encoded by putative highly expressed genes (hence called PHXPs) generally utilize low energy consuming amino acids to reduce the biosynthetic cost of the essential proteins. In our study, we found that this idea was not supported in the case of actinobacteria. With the actinobacteria, the energy costs of PHXPs varied in relation to their niche. Free-living, including aquatic, soil and extremophilic, and plant-associated actinobacteria were found to use energetically expensive amino acids in their PHXPs. An exception occurred with some animal-host-associated actinobacteria that used energy efficient amino acids. One explanation for these results may be due to the diverse metabolic patterns exhibited by actinobacteria under varied niches influenced by nutritional availability and physical environment.

Effects of varying nitrogen sources on amino acid synthesis costs in Arabidopsis thaliana under different light and carbon-source conditions.

Plants as sessile organisms cannot escape their environment and have to adapt to any changes in the availability of sunlight and nutrients. The quantification of synthesis costs of metabolites, in terms of consumed energy, is a prerequisite to understand trade-offs arising from energetic limitations. Here, we examine the energy consumption of amino acid synthesis in Arabidopsis thaliana. To quantify these costs in terms of the energy equivalent ATP, we introduce an improved cost measure based on flux balance analysis and apply it to three state-of-the-art metabolic reconstructions to ensure robust results. We present the first systematic in silico analysis of the effect of nitrogen supply (nitrate/ammonium) on individual amino acid synthesis costs as well as of the effect of photoautotrophic and heterotrophic growth conditions, integrating day/night-specific regulation. Our results identify nitrogen supply as a key determinant of amino acid costs, in agreement with experimental evidence. In addition, the association of the determined costs with experimentally observed growth patterns suggests that metabolite synthesis costs are involved in shaping regulation of plant growth. Finally, we find that simultaneous uptake of both nitrogen sources can lead to efficient utilization of energy source, which may be the result of evolutionary optimization.

Bioinformatic investigation of the cost management strategies of five oral microbes.

Some amino acids are more energetically costly to synthesize de novo, therefore many microbes have evolved to regulate the metabolic expenditure of the cell and reduce the energy burden of extracellular unrecyclable proteins. Several oral bacterial species take up amino acids and peptides obtained from proteolysis of host proteins and hence do not rely only on de novo synthesis. The aim of this study was to investigate if five oral bacterial species implement cost management strategies to reduce the energy burden of extracellular unrecyclable proteins. Since the relative de novo amino acid synthesis costs are proportional to the masses of the amino acids, the energy costs of producing proteins were assessed by calculating the mean amino acid mass for each protein. For Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, Prevotella intermedia and Streptococcus sanguinis, the outer membrane/extracellular proteins are made up of a much larger percentage of lower average mass amino acids whereas cytoplasmic proteins are made up of a larger proportion of higher average mass amino acid residues. These results are consistent with the five oral bacterial species employing energy-saving mechanisms in the production of extracellular unrecyclable proteins. Interestingly, the P. gingivalis and S. sanguinis genomes exhibited significantly lower predicted mean amino acid masses compared with those of the genomes of the other three species, suggesting that this may provide them with an energy advantage with respect to protein biosynthetic cost.

Metabolic costs of amino acid and protein production in Escherichia coli.

Escherichia coli is the most popular microorganism for the production of recombinant proteins and is gaining increasing importance for the production of low-molecular weight compounds such as amino acids. The metabolic cost associated with the production of amino acids and (recombinant) proteins from glucose, glycerol and acetate was determined using three different computational techniques to identify those amino acids that put the highest burden on the biosynthetic machinery of E. coli. Comparing the costs of individual amino acids, we find that methionine is the most expensive amino acid in terms of consumed mol of ATP per molecule produced, while leucine is the most expensive amino acid when taking into account the cellular abundances of amino acids. Moreover, we show that the biosynthesis of a large number of amino acids from glucose and particularly from glycerol provides a surplus of energy, which can be used to balance the high energetic cost of amino acid polymerization.

Genome economization in the endosymbiont of the wood roach Cryptocercus punctulatus due to drastic loss of amino acid synthesis capabilities.

Cockroaches (Blattaria: Dictyoptera) harbor the endosymbiont Blattabacterium sp. in their abdominal fat body. This endosymbiont is involved in nitrogen recycling and amino acid provision to its host. In this study, the genome of Blattabacterium sp. of Cryptocercus punctulatus (BCpu) was sequenced and compared with those of the symbionts of Blattella germanica and Periplaneta americana, BBge and BPam, respectively. The BCpu genome consists of a chromosome of 605.7 kb and a plasmid of 3.8 kb and is therefore approximately 31 kb smaller than the other two aforementioned genomes. The size reduction is due to the loss of 55 genes, 23 of which belong to biosynthetic pathways for amino acids. The pathways for the production of tryptophan, leucine, isoleucine/threonine/valine, methionine, and cysteine have been completely lost. Additionally, the genes for the enzymes catalyzing the last steps of arginine and lysine biosynthesis, argH and lysA, were found to be missing and pseudogenized, respectively. These gene losses render BCpu auxotrophic for nine amino acids more than those corresponding to BBge and BPam. BCpu has also lost capacities for sulfate reduction, production of heme groups, as well as genes for several other unlinked metabolic processes, and genes present in BBge and BPam in duplicates. Amino acids and cofactors that are not synthesized by BCpu are either produced in abundance by hindgut microbiota or are provisioned via a copious diet of dampwood colonized by putrefying microbiota, supplying host and Blattabacterium symbiont with the necessary nutrients and thus permitting genome economization of BCpu.

Amino acid biosynthetic cost and protein conservation.

Protein products of highly expressed genes tend to favor amino acids that have lower average biosynthetic costs (i.e., they exhibit metabolic efficiency). While this trend has been observed in several studies, the specific sites where cost-reducing substitutions accumulate have not been well characterized. Toward that end, weighted costs in conserved and variable positions were evaluated across a total of 9,119 homologous proteins in four mammalian orders (primate, carnivore, rodent, and artiodactyls), which together contain a total of 20,457,072 amino acids. Degree of conservation at homologous positions in these mammalian proteins and average-weighted cost across all positions within a single protein are significantly correlated. Dividing human genes into two classes (those with and those without CpG islands in their promoters) suggests that humans also preferentially utilize less costly amino acids in highly expressed genes. In contrast to the intuitive expectation that the relatively weak selective force associated with metabolic efficiency would be a selection pressure in complex multicellular organisms, the overall level of selective constraint within the variable regions of mammalian proteins allows the metabolic efficiency to derive a reduction of overall biosynthetic cost, particularly in genes with the highest levels of expression.

Diet drives convergence in gut microbiome functions across mammalian phylogeny and within humans.

Coevolution of mammals and their gut microbiota has profoundly affected their radiation into myriad habitats. We used shotgun sequencing of microbial community DNA and targeted sequencing of bacterial 16S ribosomal RNA genes to gain an understanding of how microbial communities adapt to extremes of diet. We sampled fecal DNA from 33 mammalian species and 18 humans who kept detailed diet records, and we found that the adaptation of the microbiota to diet is similar across different mammalian lineages. Functional repertoires of microbiome genes, such as those encoding carbohydrate-active enzymes and proteases, can be predicted from bacterial species assemblages. These results illustrate the value of characterizing vertebrate gut microbiomes to understand host evolutionary histories at a supraorganismal level.

Evolutionary systems biology of amino acid biosynthetic cost in yeast.

Every protein has a biosynthetic cost to the cell based on the synthesis of its constituent amino acids. In order to optimise growth and reproduction, natural selection is expected, where possible, to favour the use of proteins whose constituents are cheaper to produce, as reduced biosynthetic cost may confer a fitness advantage to the organism. Quantifying the cost of amino acid biosynthesis presents challenges, since energetic requirements may change across different cellular and environmental conditions. We developed a systems biology approach to estimate the cost of amino acid synthesis based on genome-scale metabolic models and investigated the effects of the cost of amino acid synthesis on Saccharomyces cerevisiae gene expression and protein evolution. First, we used our two new and six previously reported measures of amino acid cost in conjunction with codon usage bias, tRNA gene number and atomic composition to identify which of these factors best predict transcript and protein levels. Second, we compared amino acid cost with rates of amino acid substitution across four species in the genus Saccharomyces. Regardless of which cost measure is used, amino acid biosynthetic cost is weakly associated with transcript and protein levels. In contrast, we find that biosynthetic cost and amino acid substitution rates show a negative correlation, but for only a subset of cost measures. In the economy of the yeast cell, we find that the cost of amino acid synthesis plays a limited role in shaping transcript and protein expression levels compared to that of translational optimisation. Biosynthetic cost does, however, appear to affect rates of amino acid evolution in Saccharomyces, suggesting that expensive amino acids may only be used when they have specific structural or functional roles in protein sequences. However, as there appears to be no single currency to compute the cost of amino acid synthesis across all cellular and environmental conditions, we conclude that a systems approach is necessary to unravel the full effects of amino acid biosynthetic cost in complex biological systems.

A novel approach for determining environment-specific protein costs: the case of Arabidopsis thaliana.

MOTIVATION: Comprehensive understanding of cellular processes requires development of approaches which consider the energetic balances in the cell. The existing approaches that address this problem are based on defining energy-equivalent costs which do not include the effects of a changing environment. By incorporating these effects, one could provide a framework for integrating 'omics' data from various levels of the system in order to provide interpretations with respect to the energy state and to elicit conclusions about putative global energy-related response mechanisms in the cell. RESULTS: Here we define a cost measure for amino acid synthesis based on flux balance analysis of a genome-scale metabolic network, and develop methods for its integration with proteomics and metabolomics data. This is a first measure which accounts for the effect of different environmental conditions. We applied this approach to a genome-scale network of Arabidopsis thaliana and calculated the costs for all amino acids and proteins present in the network under light and dark conditions. Integration of function and process ontology terms in the analysis of protein abundances and their costs indicates that, during the night, the cell favors cheaper proteins compared with the light environment. However, this does not imply that there is squandering of resources during the day. The results from the association analysis between the costs, levels and well-defined expenses of amino acid synthesis, indicate that our approach not only captures the adjustment made at the switch of conditions, but also could explain the anticipation of resource usage via a global energy-related regulatory mechanism of amino acid and protein synthesis.

Do amino acid biosynthetic costs constrain protein evolution in Saccharomyces cerevisiae?

Prokaryotic organisms preferentially utilize less energetically costly amino acids in highly expressed genes. Studies have shown that the proteome of Saccharomyces cerevisiae also exhibits this behavior, but only in broad terms. This study examines the question of metabolic efficiency as a proteome-shaping force at a finer scale, examining whether trends consistent with cost minimization as an evolutionary force are present independent of protein function and amino acid physicochemical property, and consistently with respect to amino acid biosynthetic costs. Inverse correlations between the average amino acid biosynthetic cost of the protein product and the levels of gene expression in S. cerevisiae are consistent with natural selection to minimize costs. There are, however, patterns of amino acid usage that raise questions about the strength (and possibly the universality) of this selective force in shaping S. cerevisiae's proteome.

Assessment of technological options and economical feasibility for cyanophycin biopolymer and high-value amino acid production.

Major transitions can be expected within the next few decades aiming at the reduction of pollution and global warming and at energy saving measures. For these purposes, new sustainable biorefinery concepts will be needed that will replace the traditional mineral oil-based synthesis of specialty and bulk chemicals. An important group of these chemicals are those that comprise N-functionalities. Many plant components contained in biomass rest or waste stream fractions contain these N-functionalities in proteins and free amino acids that can be used as starting materials for the synthesis of biopolymers and chemicals. This paper describes the economic and technological feasibility for cyanophycin production by fermentation of the potato waste stream Protamylassetrade mark or directly in plants and its subsequent conversion to a number of N-containing bulk chemicals.

Selection on synthesis cost affects interprotein amino acid usage in all three domains of life.

Most investigations of the forces shaping protein evolution have focused on protein function. However, cells are typically 50%-75% protein by dry weight, with protein expression levels distributed over five orders of magnitude. Cells may, therefore, be under considerable selection pressure to incorporate amino acids that are cheap to synthesize into proteins that are highly expressed. Such selection pressure has been demonstrated to alter amino acid usage in a few organisms, but whether "cost selection" is a general phenomenon remains unknown. One reason for this is that reliable protein expression level data is not available for most organisms. Accordingly, I have developed a new method for detecting cost selection. This method depends solely on interprotein gradients in amino acid usage. Applying it to an analysis of 43 whole genomes from all three domains of life, I show that selection on the synthesis cost of amino acids is a pervasive force in shaping the composition of proteins. Moreover, some amino acids have different price tags for different organisms--the cost of amino acids is changed for organisms living in hydrothermal vents compared with those living at the sea surface or for organisms that have difficulty acquiring elements such as nitrogen compared with those that do not--so I also investigated whether differences between organisms in amino acid usage might reflect differences in synthesis or acquisition costs. The results suggest that organisms evolve to alter amino acid usage in response to environmental conditions.

Comparative anaerobic treatment of wastewater from pharmaceutical, brewery, paper and amino acid producing industries.

This study concerned the anaerobic treatment of five different industrial wastewaters with a diverse and complex chemical composition. The kinetics of biotransformation of this wastewater at different chemical oxygen demand (COD) were studied in a batch reactor. Wastewater from an amino acid producing industry (Fermex) and from a tank that received several types of wastewaters (collector) contained 0.83 g l(-1) and 0.085 g l(-1) sulfate, respectively. During the study period of 20 days, methane formation was observed in all types of wastewaters. Studies on COD biodegradation showed the reaction velocity was higher for Fermex wastewater and lower for collector wastewater, with values of 0.0022 h(-1) and 0.0011 h(-1), respectively. A lower methanogenic activity of 0.163 g CH4 day(-1) g(-1) volatile suspended solids (VSS) and 0.20 g CH4 day(-1) g(-1) VSS, respectively, was observed for paper producing and brewery wastewater. Adapted granular sludge showed the best biodegradation of COD during the 20-day period. The sulfate-reducing activity in pharmaceutical and collector wastewater was studied. A positive effect of sulfate-reducing activity on methanogenic activity was noted for both types of wastewaters, both of which contained sulfate ions. All reactions of methane generation for the tested industrial wastewaters were first-order. The results of this study suggest that the tested wastewaters are amenable to anaerobic treatment.

Functional genomic hypothesis generation and experimentation by a robot scientist.

The question of whether it is possible to automate the scientific process is of both great theoretical interest and increasing practical importance because, in many scientific areas, data are being generated much faster than they can be effectively analysed. We describe a physically implemented robotic system that applies techniques from artificial intelligence to carry out cycles of scientific experimentation. The system automatically originates hypotheses to explain observations, devises experiments to test these hypotheses, physically runs the experiments using a laboratory robot, interprets the results to falsify hypotheses inconsistent with the data, and then repeats the cycle. Here we apply the system to the determination of gene function using deletion mutants of yeast (Saccharomyces cerevisiae) and auxotrophic growth experiments. We built and tested a detailed logical model (involving genes, proteins and metabolites) of the aromatic amino acid synthesis pathway. In biological experiments that automatically reconstruct parts of this model, we show that an intelligent experiment selection strategy is competitive with human performance and significantly outperforms, with a cost decrease of 3-fold and 100-fold (respectively), both cheapest and random-experiment selection.

Industrial production of amino acids by coryneform bacteria.

In the 1950s Corynebacterium glutamicum was found to be a very efficient producer of L-glutamic acid. Since this time biotechnological processes with bacteria of the species Corynebacterium developed to be among the most important in terms of tonnage and economical value. L-Glutamic acid and L-lysine are bulk products nowadays. L-Valine, L-isoleucine, L-threonine, L-aspartic acid and L-alanine are among other amino acids produced by Corynebacteria. Applications range from feed to food and pharmaceutical products. The growing market for amino acids produced with Corynebacteria led to significant improvements in bioprocess and downstream technology as well as in molecular biology. During the last decade big efforts were made to increase the productivity and to decrease the production costs. This review gives an overview of the world market for amino acids produced by Corynebacteria. Significant improvements in bioprocess technology, i.e. repeated fed batch or continuous production are summarised. Bioprocess technology itself was improved furthermore by application of more sophisticated feeding and automatisation strategies. Even though several amino acids developed towards commodities in the last decade, side aspects of the production process like sterility or detection of contaminants still have increasing relevance. Finally one focus of this review is on recent developments in downstream technology.

Economic aspects of amino acids production.

Amino acids represent basic elements of proteins, which as a main source of nutrition themselves serve as a major reserve for maintaining essential functions of humans as well as animals. Taking the recent state of scientific knowledge into account, the industrial sector of amino acids is a priori "suitable" to a specific kind of an ecologically sound way of production, which is based on biotechnology. The following article may point out characteristics of this particular industrial sector and illustrates the applicability of the latest economic methods, founded on development of the discipline of bionics in order to describe economic aspects of amino acids markets. The several biochemical and technological fields of application of amino acids lead to specific market structures in high developed and permanently evolving systems. The Harvard tradition of industrial economics explains how market structures mould the behaviour of the participants and influences market results beyond that. A global increase in intensity of competition confirms the notion that the supply-side is characterised by asymmetric information in contrast to Kantzenbachs concept of "narrow oligopoly" with symmetrical shared knowledge about market information. Departing from this point, certain strategies of companies in this market form shall be derived. The importance of Research and Development increases rapidly and leads to innovative manufacturing methods which replace more polluting manufacturing processes like acid hydrolysis. In addition to these modifications within the production processes the article deals furthermore with the pricing based on product life cycle concept and introduces specific applications of tools like activity based costing and target costing to the field of amino acid production. The authors come to the conclusion that based on a good transferability of latest findings in bionics and ecological compatibility competitors in amino acids manufacturing are well advised to exercise concepts of the management of complex systems in order to choose the right strategy towards gaining market leadership.

Development of applied microbiology to modern biotechnology in Japan.

Development of modern biotechnology in Japan is characterized by unique contributions from applied microbiology and bioindustry. This review tries to summarize these original contributions with special emphasis on industrial production of useful substances by microorganisms. In the first part, development of applied microbiology and bioindustry in the last half of the twentieth century is summarized with a brief overview of the traditional background. In the second part, recent progress is reviewed with citation of typical achievements in biotechnology, applied enzymology, secondary metabolites, genetic engineering, and screening of microbial diversity, respectively.

Papain catalysed hydantoin hydrolysis in the synthesis of amino acids.

Papain has been shown, for the first time, to exhibit hydantoinase activity. It hydrolyzes the 5-mono and 5,5'-disubstituted hydantoins with linear and cyclic substituents, with a higher activity for the latter, to the corresponding N-carbamoyl amino acids, which on chemical hydrolysis yield the corresponding amino acids. The up-scaling of this simple procedure could be a major break-through for amino acid synthesis in chemical and pharmaceutical industries.