Production Of Essential Amino Acids Research Articles

Comparative transcriptomics of aphid species that diverged > 22 MYA reveals genes that are important for the maintenance of their symbiosis

Most plant-sap feeding insects have obligate relationships with maternally transmitted bacteria. Aphids require their nutritional endosymbiont, Buchnera aphidicola, for the production of essential amino acids. Such endosymbionts are harbored inside of specialized insect cells called bacteriocytes. Here, we use comparative transcriptomics of bacteriocytes between two recently diverged aphid species, Myzus persicae and Acyrthosiphon pisum, to identify key genes that are important for the maintenance of their nutritional mutualism. The majority of genes with conserved expression profiles in M. persicae and A. pisum are for orthologs previously identified in A. pisum to be important for the symbiosis. However, asparaginase which produces aspartate from asparagine was significantly up-regulated only in A. pisum bacteriocytes, potentially because Buchnera of M. persicae encodes its own asparaginase enzyme unlike Buchnera of A. pisum resulting in Buchnera of A. pisum to be dependent on its aphid host for aspartate. One-to-one orthologs that explained the most amount of variation for bacteriocyte specific mRNA expression for both species includes a collaborative gene for methionine biosynthesis, multiple transporters, a horizontally transmitted gene, and secreted proteins. Finally, we highlight species-specific gene clusters which may contribute to host adaptations and/or accommodations in gene regulation to changes in the symbiont or the symbiosis.

Host’s demand for essential amino acids is compensated by an extracellular bacterial symbiont in a hemipteran insect model

Plant sap is a nutritionally unbalanced diet that constitutes a challenge for insects that feed exclusively on it. Sap-sucking hemipteran insects generally overcome this challenge by harboring beneficial microorganisms in their specialized symbiotic organ, either intracellularly or extracellularly. Genomic information of these bacterial symbionts suggests that their primary role is to supply essential amino acids, but empirical evidence has been virtually limited to the intracellular symbiosis between aphids and Buchnera. Here we investigated the amino acid complementation by the extracellular symbiotic bacterium Ishikawaella harbored in the midgut symbiotic organ of the stinkbug Megacopta punctatissima. We evaluated amino acid compositions of the phloem sap of plants on which the insect feeds, as well as those of its hemolymph, whole body hydrolysate, and excreta. The results highlighted that the essential amino acids in the diet are apparently insufficient for the stinkbug development. Experimental symbiont removal caused severe shortfalls of some essential amino acids, including branched-chain and aromatic amino acids. In vitro culturing of the isolated symbiotic organ demonstrated that hemolymph-circulating metabolites, glutamine and trehalose, efficiently fuel the production of essential amino acids. Branched-chain amino acids and aromatic amino acids are the ones preferentially synthesized despite the symbiont’s synthetic capability of all essential amino acids. These results indicate that the symbiont-mediated amino acid compensation is quantitatively optimized in the stinkbug-Ishikawaella gut symbiotic association as in the aphid-Buchnera intracellular symbiotic association. The convergence of symbiont functions across distinct nutritional symbiotic systems provides insight into how host-symbiont interactions have been shaped over evolutionary time.

Antibiotics in the soil and their effect on the soil microbiot

Antibiotics have been crucial in the fight against infectious diseases for the past 50 years. In agriculture they are widely used in the treatment of animals, birds and aquaculture, to prevent spoilage of feed, as stimulators of growth and productivity of livestock, in the production of essential amino acids as impurities in feed, and so on. At present, the use of antibiotics in animal husbandry has become excessive due to the prevention of global epidemics. In turn, the ingress of antibiotics into water and soil, in particular through organic fertilizers, poses a potential threat to these environments. Thus, a variety of antibiotic resistance genes (GRAs) are spreading in soil microorganisms, which is currently a global health problem. It is believed that the stability of antibiotics after entering the soil is mainly due to their rate of decomposition and sorption to the organic soil matrix. A wide range of values of the half-life (DT50) of these compounds in soils indicates that their stability depends on a number of factors: soil properties, climatic conditions (temperature, precipitation, and humidity), physicochemical characteristics of antibiotics. High antimicrobial activity of antibiotics in the soil differentially inhibits the development of soil microorganisms, affects their species composition, which can cause changes in the ecological functionality of the soil. Thus, even low concentrations of antibiotics significantly reduce soil respiration. This phenomenon is especially noticeable in the presence of sulfamethoxazole, sulfamethazine, sulfadiazine and trimethoprim in the soil. The presence of antibiotics in the soil affects the processes of nitrification and / or denitrification, and the inhibition of these processes depends on the duration of exposure and the type of compound. Monensin and chlortetracycline at concentrations of 0.01–0.1 and 0.0003–0.3 mg/kg of soil do not affect nitrification at all. Antibiotics also affect the rate of iron transformation in the soil. Thus, sulfadiazine and monensin block the reduction of iron (Fe (III)) in the soil from a few days to 50 days. It should be noted that the lack of standardized tests hinders research that would lead to generalized conclusions about the effects of antibiotics on biogeochemical cycles, in particular on iron circulation. An important indicator of the response to antibiotics in the soil is considered to be the change in the enzymatic activity of dehydrogenase, phosphatase and urease of soil microorganisms, which may be associated with growth inhibition or death of sensitive microorganisms. In addition, the presence of some antibiotics in the soil can cause over-population of fungal populations, which are generally less sensitive to antibiotics than bacteria. There is evidence that antibiotics alter the enzymatic activity of soil microorganisms, especially they affect the ability to metabolize carbon of various origins. In addition, antibiotics not only affect the total number of microbiota, but also the relative content of different groups (gram-negative and gram-positive bacteria, fungi) in microbial populations. The importance of GRA studies of soil microorganisms is that they have led to the discovery of new genes responsible for bacterial resistance to antibiotics.

Evolutionarily recent dual obligatory symbiosis among adelgids indicates a transition between fungus- and insect-associated lifestyles

Adelgids (Insecta: Hemiptera: Adelgidae) form a small group of insects but harbor a surprisingly diverse set of bacteriocyte-associated endosymbionts, which suggest multiple replacement and acquisition of symbionts over evolutionary time. Specific pairs of symbionts have been associated with adelgid lineages specialized on different secondary host conifers. Using a metagenomic approach, we investigated the symbiosis of the Adelges laricis/Adelgestardus species complex containing betaproteobacterial (“Candidatus Vallotia tarda”) and gammaproteobacterial (“Candidatus Profftia tarda”) symbionts. Genomic characteristics and metabolic pathway reconstructions revealed that Vallotia and Profftia are evolutionary young endosymbionts, which complement each other’s role in essential amino acid production. Phylogenomic analyses and a high level of genomic synteny indicate an origin of the betaproteobacterial symbiont from endosymbionts of Rhizopus fungi. This evolutionary transition was accompanied with substantial loss of functions related to transcription regulation, secondary metabolite production, bacterial defense mechanisms, host infection, and manipulation. The transition from fungus to insect endosymbionts extends our current framework about evolutionary trajectories of host-associated microbes.

Capacidad metabólica de cultivos mixtos probióticos formados por cepas de Lactobacillus y Bifidobacterium para uso en alimentos lácteos fermentados funcionales

The metabolic capacity of probiotic mixed cultures formed by Lactobacillus and Bifidobacterium strains was assessed through the determination of the acidification profile and the production of amino acids and volatile compounds, during the fermentation of ultra-pasteurized skim milk. Two mixed cultures formed by [L. acidophilus + B. bifidum] and [L. acidophilus + B. animalis] stood out in the production of essential amino acids: His, Ile, Leu, Met, Thr, Trp and Val. As well as increased production of volatile compounds such as: acetoin, 2-heptanone, 2-nonanone, acetic acid, acid butanoic and hexanoic acid, in contrast to commercial fermented dairy products. Additionally, these bacterial mixed cultures were characterized by the production of distinctive volatile compounds: 1-heptanol, diisobutylcarninol, hemellitol, 1-dodecanol, acetone, 2-pentanone, 2-undecanone, ethyl acetate, benzaldehyde as well as valeric, acetyl valeric and isopropylpyruvic acids. Finally, the culture formed by [L. acidophilus + B. bifidum] presented a good acidification profile with a lactic acid production of 26.1 ± 0.1 g / L and pH 3.6 at the end of fermentation. This data suggests a great potential of these mixed cultures to improve the nutritional value and organoleptic characteristics of fermented dairy products, when added as starter or adjunct cultures in the fermentation process.

Indian oil sardine (Sardinella longiceps) gut derived Bacillus safensis SDG14 with enhanced probiotic competence for food and feed applications

Probiotics are considered as functional food as they provide health benefits along with traditional nutrition. Spore forming probiotic Bacillus are of commercial interest than Lactic Acid Bacillus due to their relatively lower cost of production and higher survivability. In the present study we identified the bacterial strain SDG14 isolated from Indian oil Sardine by Average Nucleotide Identity of whole genome sequence. The whole genome of SDG14 was also explored for pathogenicity, the presence of genes responsible for probiotic traits such as spore formation, resistance to host gastrointestinal tract conditions, adhesion to intestinal mucosa, interference in pathogen survival, expression of bacteriocins, oxidative and other stress responses, absorption of nutrition, production of essential amino acids and vitamins. Wet lab experiments for probiotic characterization were also conducted. The organism was confirmed to be Bacillus safensis SDG14. The possible pathogenicity of the organism was also ruled out by in silico analysis. Bacillus safensis SDG14 was able to survive at pH 3 and bile salt concentration of 0.5% (w/v). The adhesion index of Bacillus safensis SDG14 on HEp-2 was 36.82 ± 5.93 and 45.54 ± 9.55 respectively after 60 and 90 min of incubation and self aggregation percentage was 18.4 ± 0.48% after 3 h. Bacillus safensis SDG14 produced bacteriocin and co-aggregated with E. coli, Salmonella Typhimurium and Pseudomonas aeruginosa. The genomic data supported the findings of wet lab study and vice versa. Bacillus safensis SDG14 was proved to be a non-pathogenic, spore forming, pH and bile salt resistant, bacteriocin, amino acid and vitamin producing probiotic with proposed food and feed applications.

Evidence for Succession and Putative Metabolic Roles of Fungi and Bacteria in the Farming Mutualism of the Ambrosia Beetle Xyleborus affinis.

The bacterial and fungal community involved in ambrosia beetle fungiculture remains poorly studied compared to the famous fungus-farming ants and termites. Here we studied microbial community dynamics of laboratory nests, adults, and brood during the life cycle of the sugarcane shot hole borer, Xyleborus affinis We identified a total of 40 fungal and 428 bacterial operational taxonomic units (OTUs), from which only five fungi (a Raffaelea fungus and four ascomycete yeasts) and four bacterial genera (Stenotrophomonas, Enterobacter, Burkholderia, and Ochrobactrum) can be considered the core community playing the most relevant symbiotic role. Both the fungal and bacterial populations varied significantly during the beetle's life cycle. While the ascomycete yeasts were the main colonizers of the gallery early on, the Raffaelea and other filamentous fungi appeared after day 10, at the time when larval hatching happened. Regarding bacteria, Stenotrophomonas and Enterobacter dominated overall but decreased in foundresses and brood with age. Finally, inferred analyses of the putative metabolic capabilities of the bacterial microbiome revealed that they are involved in (i) degradation of fungal and plant polymers, (ii) fixation of atmospheric nitrogen, and (iii) essential amino acid, cofactor, and vitamin provisioning. Overall, our results suggest that yeasts and bacteria are more strongly involved in supporting the beetle-fungus farming symbiosis than previously thought.IMPORTANCE Ambrosia beetles farm their own food fungi within tunnel systems in wood and are among the three insect lineages performing agriculture (the others are fungus-farming ants and termites). In ambrosia beetles, primary ambrosia fungus cultivars have been regarded essential, whereas other microbes have been more or less ignored. Our KEGG analyses suggest so far unknown roles of yeasts and bacterial symbionts, by preparing the tunnel walls for the primary ambrosia fungi. This preparation includes enzymatic degradation of wood, essential amino acid production, and nitrogen fixation. The latter is especially exciting because if it turns out to be present in vivo in ambrosia beetles, all farming animals (including humans) are dependent on atmospheric nitrogen fertilization of their crops. As previous internal transcribed spacer (ITS) metabarcoding approaches failed on covering the primary ambrosia fungi, our 18S metabarcoding approach can also serve as a template for future studies on the ambrosia beetle-fungus symbiosis.

Key Transport and Ammonia Recycling Genes Involved in Aphid Symbiosis Respond to Host-Plant Specialization.

Microbes are known to influence insect-plant interactions; however, it is unclear if host-plant diet influences the regulation of nutritional insect symbioses. The pea aphid, Acyrthosiphon pisum, requires its nutritional endosymbiont, Buchnera, for the production of essential amino acids. We hypothesize that key aphid genes that regulate the nutritional symbioses respond to host-plant diet when aphids feed on a specialized (alfalfa) compared to a universal host-plant diet (fava), which vary in amino acid profiles. Using RNA-Seq and whole genome bisulfite sequencing, we measured gene expression and DNA methylation profiles for such genes when aphids fed on either their specialized or universal host-plant diets. Our results reveal that when aphids feed on their specialized host-plant they significantly up-regulate and/or hypo-methylate key aphid genes in bacteriocytes related to the amino acid metabolism, including glutamine synthetase in the GOGAT cycle that recycles ammonia into glutamine and the glutamine transporter ApGLNT1. Moreover, regardless of what host-plant aphids feed on we observed significant up-regulation and differential methylation of key genes involved in the amino acid metabolism and the glycine/serine metabolism, a metabolic program observed in proliferating cancer cells potentially to combat oxidative stress. Based on our results, we suggest that this regulatory response of key symbiosis genes in bacteriocytes allows aphids to feed on a suboptimal host-plant that they specialize on.

Cooperative Metabolism in a Three-Partner Insect-Bacterial Symbiosis Revealed by Metabolic Modeling

ABSTRACTAn important factor determining the impact of microbial symbionts on their animal hosts is the balance between the cost of nutrients consumed by the symbionts and the benefit of nutrients released back to the host, but the quantitative significance of nutrient exchange in symbioses involving multiple microbial partners has rarely been addressed. In this study on the association between two intracellular bacterial symbionts, “Candidatus Portiera aleyrodidarum” and “Candidatus Hamiltonella defensa,” and their animal host, the whitefly Bemisia tabaci, we apply metabolic modeling to investigate host-symbiont nutrient exchange. Our in silico analysis revealed that >60% of the essential amino acids and related metabolites synthesized by “Candidatus Portiera aleyrodidarum” are utilized by the host, including a substantial contribution of nitrogen recycled from host nitrogenous waste, and that these interactions are required for host growth. In contrast, “Candidatus Hamiltonella defensa” retains most or all of the essential amino acids and B vitamins that it is capable of synthesizing. Furthermore, “Candidatus Hamiltonella defensa” suppresses host growth in silico by competition with “Candidatus Portiera aleyrodidarum” for multiple host nutrients, by suppressing “Candidatus Portiera aleyrodidarum” growth and metabolic function, and also by consumption of host nutrients that would otherwise be allocated to host growth. The interpretation from these modeling outputs that “Candidatus Hamiltonella defensa” is a nutritional parasite could not be inferred reliably from gene content alone but requires consideration of constraints imposed by the structure of the metabolic network. Furthermore, these quantitative models offer precise predictions for future experimental study and the opportunity to compare the functional organization of metabolic networks in different symbioses.IMPORTANCE The metabolic functions of unculturable intracellular bacteria with much reduced genomes are traditionally inferred from gene content without consideration of how the structure of the metabolic network may influence flux through metabolic reactions. The three-compartment model of metabolic flux between two bacterial symbionts and their insect host constructed in this study revealed that one symbiont is structured to overproduce essential amino acids for the benefit of the host, but the essential amino acid production in the second symbiont is quantitatively constrained by the structure of its network, rendering it “selfish” with respect to these nutrients. This study demonstrates the importance of quantitative flux data for elucidation of the metabolic function of symbionts. The in silico methodology can be applied to other symbioses with intracellular bacteria.

Effects of culture substrates on taste component content and taste quality of Lentinula edodes

SummaryIn this research, the effects of culture substrates on taste component content of Lentinula edodes were studied, and the resulting taste quality of L. edodes was evaluated. The results revealed that single‐carbon and single‐nitrogen sources were beneficial to production of soluble sugars and polyols, organic acids and sweet amino acids, whereas a single‐carbon source was beneficial to essential amino acid production, and a mixed‐carbon source was beneficial to umami 5′‐nucleotides and high mushroom production yield. High C/N values were beneficial to trehalose, arabitol, malic acid, and succinic acid production, while low C/N values were beneficial to mannitol and citric acid production. L. edodes fruiting bodies, harvested from a culture substrate containing single carbon, high proportion of cereal bran, had a more palatable taste quality, while a substrate containing bagasse and low C/N values was not suitable for L. edodes cultivation due to unfavourable taste quality. These results provide cultivation information how to obtain fruiting bodies L. edodes with more taste components.

Inoculation of abscisic acid-producing endophytic bacteria enhances salinity stress tolerance in Oryza sativa

Salinity hinders plant growth and results in reduced crop yield. The use of plant growth-promoting endophytic bacteria is an eco-friendly strategy to counteract such stresses and confer tolerance to the host. Endophytic bacteria have been recognized for their active role in auxin production; however, little is known about their ability to produce abscisic acid (ABA). In recent studies, the bacterial endophyte Bacillus amyloliquefaciens RWL-1 has been found to produce ABA, and as such, has the potential to increase plant resistance to salinity stress. Results showed that RWL-1 produced varying concentrations of ABA (0.32±0.015–0.14±0.030ngmL−1) under normal and saline conditions. The ability of RWL-1 to produce ABA was reduced in response to increasing salinity; however, it maintained its growth by up-regulating production of essential amino acids (glutamic acid and proline). To further investigate the potential of this endophytic bacterium, a plant-microbe interaction experiment was conducted which showed that RWL-1 inoculation significantly increased growth attributes of rice plants as compared to non-inoculated control plants under salinity stress. Micrographs also revealed active symbiosis of RWL-1 with plant roots under normal and salinity stress conditions. The essential amino acids (glutamic acid, aspartic acid, phenylalanine, proline, and cysteine) were significantly up-regulated by RWL-1 inoculation under salinity stress. In addition, the stress-sensitive endogenous ABA levels were significantly reduced, whereas the levels of endogenous salicylic acid were significantly higher in RWl-1-inoculated plants than in control plants exposed to the same level of salinity stress. The current findings suggest that the phytohormone-producing abilities of endophytic bacteria can increase plant resistance to salinity, in turn improving agricultural productivity.

Matching the supply of bacterial nutrients to the nutritional demand of the animal host.

Various animals derive nutrients from symbiotic microorganisms with much-reduced genomes, but it is unknown whether, and how, the supply of these nutrients is regulated. Here, we demonstrate that the production of essential amino acids (EAAs) by the bacterium Buchnera aphidicola in the pea aphid Acyrthosiphon pisum is elevated when aphids are reared on diets from which that EAA are omitted, demonstrating that Buchnera scale EAA production to host demand. Quantitative proteomics of bacteriocytes (host cells bearing Buchnera) revealed that these metabolic changes are not accompanied by significant change in Buchnera or host proteins, suggesting that EAA production is regulated post-translationally. Bacteriocytes in aphids reared on diet lacking the EAA methionine had elevated concentrations of both methionine and the precursor cystathionine, indicating that methionine production is promoted by precursor supply and is not subject to feedback inhibition by methionine. Furthermore, methionine production by isolated Buchnera increased with increasing cystathionine concentration. We propose that Buchnera metabolism is poised for EAA production at certain maximal rates, and the realized release rate is determined by precursor supply from the host. The incidence of host regulation of symbiont nutritional function via supply of key nutritional inputs in other symbioses remains to be investigated.

Bacteriocyte dynamics during development of a holometabolous insect, the carpenter ant Camponotus floridanus

BackgroundThe carpenter ant Camponotus floridanus harbors obligate intracellular mutualistic bacteria (Blochmannia floridanus) in specialized cells, the bacteriocytes, intercalated in their midgut tissue. The diffuse distribution of bacteriocytes over the midgut tissue is in contrast to many other insects carrying endosymbionts in specialized tissues which are often connected to the midgut but form a distinct organ, the bacteriome. C. floridanus is a holometabolous insect which undergoes a complete metamorphosis. During pupal stages a complete restructuring of the inner organs including the digestive tract takes place. So far, nothing was known about maintenance of endosymbionts during this life stage of a holometabolous insect. It was shown previously that the number of Blochmannia increases strongly during metamorphosis. This implicates an important function of Blochmannia in this developmental phase during which the animals are metabolically very active but do not have access to external food resources. Previous experiments have shown a nutritional contribution of the bacteria to host metabolism by production of essential amino acids and urease-mediated nitrogen recycling. In adult hosts the symbiosis appears to degenerate with increasing age of the animals.ResultsWe investigated the distribution and dynamics of endosymbiotic bacteria and bacteriocytes at different stages during development of the animals from larva to imago by confocal laser scanning microscopy. The number of bacteriocytes in relation to symbiont-free midgut cells varied strongly over different developmental stages. Especially during metamorphosis the relative number of bacteria-filled bacteriocytes increased strongly when the larval midgut epithelium is shed. During this developmental stage the midgut itself became a huge symbiotic organ consisting almost exclusively of cells harboring bacteria. In fact, during this phase some bacteria were also found in midgut cells other than bacteriocytes indicating a cell-invasive capacity of Blochmannia. In adult animals the number of bacteriocytes generally decreased.ConclusionsDuring the life cycle of the animals the distribution of bacteriocytes and of Blochmannia endosymbionts is remarkably dynamic. Our data show how the endosymbiont is retained within the midgut tissue during metamorphosis thereby ensuring the maintenance of the intracellular endosymbiosis despite a massive reorganization of the midgut tissue. The transformation of the entire midgut into a symbiotic organ during pupal stages underscores the important role of Blochmannia for its host in particular during metamorphosis.

Limited Endosymbiont Variation in <I>Diuraphis noxia</I> (Hemiptera: Aphididae) Biotypes From the United States and South Africa

Symbiosis allows an insect access to imbalanced food sources on which other organisms cannot survive. A bacterial endosymbiont, Buchnera aphidicola, gives aphids the ability to feed on phloem depleted of certain essential amino acids by producing those required. Pseudogenes and lower plasmid copy numbers of essential amino acid genes in B. aphidicola, endosymbiont of the Russian wheat aphid, Diuraphis noxia (Kurdjumov) (Hemiptera: Aphididae), suggest that this symbiotic relationship is degenerating. The complete endosymbiont assemblages, copy numbers of plasmids (important in essential amino acid production), and sequence variation in B. aphidicola, from 10 Russian wheat aphid biotypes, were investigated. B. aphidicola was found to be monosymbiotic in the Russian wheat aphid biotypes and other Diuraphis species examined. An insert, occurring in an inverted repeat region on the leucine plasmid, was the only variation found in the approximately 10-kb B. aphidicola sequence analyzed from each Russian wheat aphid biotype. This inverted repeat was shown previously to be conserved within the family Aphididae. The insert occurred in B. aphidicola sequences isolated from four Russian wheat aphid biotypes. Copy numbers of the leucine plasmid differ between the South African and U.S. biotypes and were similar to previously reported values for biotypes from the same geographic regions. These results suggest that B. aphidicola may still contribute to Russian wheat aphid fitness when the aphid feeds on a variety of hosts.

Integrated Metabonomic−Proteomic Analysis of an Insect−Bacterial Symbiotic System

The health of animals, including humans, is dependent on their resident microbiota, but the complexity of the microbial communities makes these associations difficult to study in most animals. Exceptionally, the microbiology of the pea aphid Acyrthosiphon pisum is dominated by a single bacterium Buchnera aphidicola (B. aphidicola). A (1)H NMR-based metabonomic strategy was applied to investigate metabolic profiles of aphids fed on a low essential amino acid diet and treated by antibiotic to eliminate B. aphidicola. In addition, differential gel electrophoresis (DIGE) with mass spectrometry was utilized to determine the alterations of proteins induced by these treatments. We found that these perturbations resulted in significant changes to the abundance of 15 metabolites and 238 proteins. Ten (67%) of the metabolites with altered abundance were amino acids, with nonessential amino acids increased and essential amino acids decreased by both perturbations. Over-represented proteins in the perturbed treatments included catabolic enzymes with roles in amino acid degradation and glycolysis, various cuticular proteins, and a C-type lectin and regucalcin with candidate defensive roles. This analysis demonstrates the central role of essential amino acid production in the relationship and identifies candidate proteins and processes underpinning the function and persistence of the association.

Production of essential amino acids from glutamate by mycetocyte symbionts of the pea aphid, Acyrthosiphon pisum

Glutamine was the most abundant amino acid constituent in the hemolymph of pea aphids, Acyrthosiphon pisum. While mycetocytes isolated from the aphids took up glutamine actively, the intracellular symbionts isolated from the mycetocytes scarcely took up the amino acid, and instead took up glutamic acid actively. [U- 14C]Glutamine incorporated by mycetocytes was converted into glutamate. When [ϵ- 15N]glutamine was introduced into mycetocytes, [ 15N]glutamate was found in the cytosol. These results suggested that glutamine taken up by mycetocytes was hydrolyzed into glutamic acid and ammonia, and at least a portion of ammonia was assimilated into glutamate in the cytosol, probably through reaction with α-ketoglutarate. When isolated symbionts were incubated with [ 15N]glutamic acid, the following amino acids were found highly labeled: alanine, aspartic acid, glutamine, isoleucine, leucine, phenylalanine, proline and valine.

Medical applications of artificial cells in transfusion, phenylketonuria, essential amino acid production, and liver support.

Etude de la modification de l'hemoglobine afin d'obtenir un substitut dans les transfusions sanguines, du remplacement d'enzyme dans les maladies metaboliques telles que la phenylcetonurie, de la bioconversion des metabolites de dechets (uree, ammoniac) en acides amines essentiels et de l'immobilisation de cultures cellulaires par des cellules artificielles

Quantitative and qualitative investigations on the seed proteins of mutants and recombinants of Pisum sativum

The proteins of the seed flour of 26 X-ray induced mutants and 5 recombinants of the species Pisum sativum were analysed quantitatively and qualitatively. The values obtained were related to the seed yield of the genotypes In this way, the protein yield as well as the production of specific amino acids per genotype were determined. The following results were obtained. 1. The mean values for the character "total protein content of the seed flour" of the genotypes studied varied between 14 and 23%. The fasciated mutant 489C produced 18-23% more seed proteins than the initial line in three subsequent generations. The mean values of some other mutants and recombinants were 10-17% higher, the lowest value being 27% lower than that of the control values. In the material investigated there is no correlation between seed size and seed protein content. 2. The buffer-soluble seed proteins of 55 mutants were electrophoretically subdivided in different subfractions and the protein patterns were determined. They are extraordinarily variable. Distinct groups of mutants do not only differ from the initial line but also from one another with regard to the number, position and breadth of their bands. A correlation between the degree of morphological deviations of specific genotypes and the composition of their seed proteins was not observed. Moreover, differences in the concentration of specific protein fractions between the genotypes were densitometrically ascertained. 3. The globulins and albumins of some genotypes were quantitatively determined and electrophoretically subdivided into subfractions which differ between different mutants with regard to their number as well as their concentration. This is especially valid for the albumins which could be essentially stronger subdivided than the globulins. 4. The amino acid spectra of all the mutants investigated agree qualitatively with the spectrum of the initial line, however, clear quantitative differences in distinct amino acids were observed. The proportion of the essential amino acids is increased in two mutants by 5 and 20% in relation to the control line. 5. The protein production of the fasciated mutant 489C was 20-70% higher than the corresponding values of the initial line in three subsequent generations regarding its high seed production. The protein yield of the early flowering mutant 46C was about 20% lower and that of the bifurcated mutant 1201A 12-31% higher as related to the initial line. Mutant 1201A shows an equally favourable situation with regard to the total content of the essential amino acids. Its lysine production exceeded the control values by 20-40%. 6. The different components of the protein synthesis can negatively be influenced by the co-operation of the mutant genes. The protein production per plant, the total production of essential amino acids as well as the lysine production are lower in the recombinants 68C/176A and 68C/1201A as compared to the parental mutants. The combination of genes 68C and 46C does not show any negative interactions.

Leaf protein concentrates: a comparison of protein production per acre of forage with that from seed and animal crops

The essential amino acid production for ten crops harvested for forage and fifteen crops harvested for seed was calculated from average crop yields in the United States for the ten-year period of 1953 to 1962. Highest yields per acre of essential amino acids were calculated from forages which could be processed into leaf protein concentrates. Alfalfa produced the highest yield per acre of essential amino acids of the twenty major crops; soybean seed was second. The calculated yields of edible protein and essential amino acids of leaf protein from corn were equal to or greater than the protein from the corn seed and several times that from animals fed the corn grain. At the present time, leaf protein concentrate may not be able to compete in cost with the by-product protein concentrates such as soybean meal, cottonseed meal, peanut meal, linseed meal, or tankage for feeding farm animals. However, further study of the production of leaf protein concentrates should be made to assure protein supplies for those regions where other adequate protein sources are lacking.