Studies on the Metabolism of Aspergillii

Endogenous respiration of Aspergillus sojae K.S. was studied in terms of biochemical analysis. It was found that the different kind of substrates was utilized for the endogenous respiratoin according to C:N ratio of the agar medium on which the mold was grown. In the mycelial mats grown on the medium of low C:N value, pool amino acids, protein, and nucleic acids were mainly utilized from the beginning while carbohydrate or lipid displayed a minor role. The corresponding amount of ammonia was formed. On the other hand, in the mycelial mats grown on the medium of rather high C:N value, carbohydrate or lipid was the major substrate of endogenous respiration in the early stages of incubation. The utilization of the nitrogenous materials and the accompanying formation of ammonia got to start only after the lapse of several hours of incubation.     

Studies on the Metabolism of Aspergilli

This report is concerned with the time course of changes of several enzymes during endogenous respiration. It was observed that enzymes such as deaminases, amidases and transaminases found in the mycelia increased in activity more or less during endogenous respiration. It was assumed that enzyme formation occurred as the result of glucose starvation or depression of carbohydrate metabolism during endogenous respiration of the mold mycelia on buffer solution.     

Note on study of the sporulation of fungi: endotrophic sporulation in the genusPenicillium

Conidiophore formation and sporulation can be induced inPenicillium sp. strain P 17 by an environmental factor—carbohydrate (carbon) starvation. Both surface and submerged mycelium, when transferred from synthetic medium to glucose-free salt solution, form conidiophores and sporulate, while in the control cultures on complete medium, vegetative growth continues. The time required for the formation of conidiophores, i.e. the induction interval, is 7–14 h and its length increases with the age of both surface and submerged mycelia. During the induction phase the mycelium undergoes autolysis, associated with degradation of energy motabolism involving the comsumption of reserve substances, a rapid drop in endogenous respiration and the endogenous reducing activity of the mycelium, a decrease in the labile phosphate concentration, proteolysis, an increase in the ammonia and orthopsphate concentration and exhaustion of readily oxidized amino acids from the pool. A transient increase in respiration occurs before differentiation of the conidiophores starts. During the second half of the induction phase, polyphenol substances and polyphenol oxidase appear in the mycelium.The enzyme is not induced by exogenous phenols. Its possible role in the sporulation of fungi is considered.     

Manganese deficiency leads to elevated amino acid pools in citric acid accumulating Aspergillus niger

Free amino acid pools have been investigated in a citric acid accumulating strain of Aspergillus niger during batch growth under manganese sufficient and deficient conditions by means of an improved chromatographic method. Studies on the mycelial content of several nitrogenous compounds under manganese sufficient and deficient conditions showed that manganese deficiency resulted in lower amino acid pool sizes during trophophase and considerable accumulation during idiophase, and in a reduction of the protein and nucleic acid contents. Addition of cycloheximide to mycelia grown with sufficient manganese also caused an elevation of free amino acid pool sizes, thus indicating that impairment of protein synthesis by manganese deficiency is responsible for the observed rise in amino acid concentration. Furthermore it was observed that the manganese deficient mycelia excreted high amounts of all amino acids suggesting that manganese deficiency may also affect membrane permeability.     

香菇复配氨基酸培养基优化及其对香菇菌丝生长生理的影响

[背景]L-氨基酸能够提供香菇生长发育的部分养分需求,利用外源添加氨基酸的方式对香菇生长基质进行优化,对于香菇产业的增产提质具有积极的发展意义.[目的]利用外源添加丙氨酸(Ala)、丝氨酸(Ser)和天冬酰胺(Asn)的方式改善培养基成分,并探究复配氨基酸促进香菇生长的可能代谢途径.[方法]在三因素三水平上,通过响应面...     

黄伞菌丝蛋白质营养价值评价

本文测定了黄伞Pholiotaadiposa菌丝粗蛋白含量和氨基酸组成,采用国际上通用的营养价值评价方法,对其蛋白质的营养价值进行全面评价,并与营养价值较高的白灵侧耳Pleurotusnebrodensis、刺芹侧耳Pleurotuseryngii和金顶侧耳Pleurotrscitrinopileatus进行比较。分析结果表明,黄伞菌丝的必需氨基酸含量最高,占其氨基酸总量的43.9%,蛋白质的氨基酸评分(AAS)、化学评分(CS)、必需氨基酸指数(EAAI)、生物价(BV)、营养指数(NI)和氨基酸比值系数分(SRCAA)分别为92.0、71.3、88.5、84.8、40.4和78.8,六项指标均比参比食用菌高。结果说明黄伞菌丝具有很高的营养价值。     

Amino Acid Metabolic Origin as an Evolutionary Influence on Protein Sequence in Yeast

The metabolic cycle of Saccharomyces cerevisiae consists of alternating oxidative (respiration) and reductive (glycolysis) energy-yielding reactions. The intracellular concentrations of amino acid precursors generated by these reactions oscillate accordingly, attaining maximal concentration during the middle of their respective yeast metabolic cycle phases. Typically, the amino acids themselves are most abundant at the end of their precursor’s phase. We show that this metabolic cycling has likely biased the amino acid composition of proteins across the S. cerevisiae genome. In particular, we observed that the metabolic source of amino acids is the single most important source of variation in the amino acid compositions of functionally related proteins and that this signal appears only in (facultative) organisms using both oxidative and reductive metabolism. Periodically expressed proteins are enriched for amino acids generated in the preceding phase of the metabolic cycle. Proteins expressed during the oxidative phase contain more glycolysis-derived amino acids, whereas proteins expressed during the reductive phase contain more respiration-derived amino acids. Rare amino acids (e.g., tryptophan) are greatly overrepresented or underrepresented, relative to the proteomic average, in periodically expressed proteins, whereas common amino acids vary by a few percent. Genome-wide, we infer that 20,000 to 60,000 residues have been modified by this previously unappreciated pressure. This trend is strongest in ancient proteins, suggesting that oscillating endogenous amino acid availability exerted genome-wide selective pressure on protein sequences across evolutionary time. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Benjamin L. de Bivort and Ethan O. Perlstein have contributed equally to this work.     

Some Nutritional and Physiological Factors Affecting the Urinary Excretion of Acid Soluble Peptides in Rats and Women

Urinary excretion of acid soluble peptide (ASP)-form amino acids was lower in rats deprived of protein than in rats fed on a 20% casein or 20% gluten diet. However, the amino acid pattern of urinary ASP was similar among each of the three dietary groups, suggesting that urinary ASP is mainly endogenous origin under these nutritional conditions.

College women who were given a meat-free protein diet for 3 days after 10 days’ protein deprivation excreted 1.4 times the amount of ASP-form amino acids during protein deprivation.

The rate of urinary excretion of ASP-form amino acids in the state of protein deprivation was proportional to the metabolic body size of organisms as far as rats and women were concerned.

Streptozotocin-induced diabetic rats excreted two times the amount of ASP-form amino acids compared with normal rats. This suggests that endogenous protein catabolism doubled in diabetic rats.

When labelled urinary ASP was injected into rats, approximately 40% of the label was recovered as urinary ASP within 24 hr. This excretion rate was far higher than that after the injection of free leucine.

The rate of urinary excretion of ASP-form amino acids correlated with that of N^τ-methylhistidine in rats.

These results favor the hypothesis that urinary ASP reflects the catabolism of body proteins.     

Comparative Physiology of Respiration in Aquatic Fungi II. The Saprolegniales, Especially Aphanomyces astaci

The endogenous respiration of six members of Saprolegniaceae (Oomycetes), Saprotegnia sp., Thraustotheca sp., Achlya sp., Dichtyuchus sp., Aphanomyces euteiches, and A. astaci were studied in the presence and in the absence of exogenous substrate using conventional manometric techniques. Glucose stimulated the rate of oxygen uptake of unstarved mycelia to some extent in all the fungi. Attempts to increase the weak stimulation of respiration by glucose in A. astaci were not successful. The respiratory quotients of the fungi tested were usually in the range of 0.7 to 0.9 during endogenous respiration, and addition of glucose increased these values more than expected. L-leucine and L-glutamic acid stimulated respiration of A. astaci only when the fugus was starved, and acetic acid and butyric acid were inhibitory. Fructose and acetic acid increased respiration in starved mycelium of A. euteiches while L-leucine and L-glutamic acid had little effect. Antimycin A, HOQNO, HCN, and fluoroacetate strongly inhibited endogenous oxygen uptake by A. astaci. Amytal and azide were also markedly inhibitory while rotenone and CO had little effect. DNP and diphenylamine inhibited respiration at a high concentration but at a lower concentration DNP was stimulatory. In contrast the respiration of Saprolegnia sp. was resistant to cyanide, antimycin A, and HOQNO. Spectrophotometric observations on homogenized mycelia of Saprolegnia sp. and of A. astaci indicated the presence of cytochrome c (551 nm), two b-type cytochromes (557 and 564 nm) and cytochrome a-a₃ (605 nm) all in approximately equimolar concentrations. In both strains CO combined with cytochrome a₃ and an unidentified pigment. A remarkable similarity in the cytochrome system seems to exist between these two strains and some members of the Leptomitales.     

Effect of Fatty Acids and Methyl Octanoate on Resting Mycelium of Boletus variegatus

The endogenous respiration of resting, submerged grown Boletus variegatus mycelium has been determined. In young cultures the intensity of the endogenous oxygen uptake was subject to great variations during the first few hours of starvation. However, by using six to eight days old mycelium the Qo₂ values could be kept at a relatively low and constant level for at least nine hours. Inhibition of the endogenous respiration was found after addition of n-saturated C-2 to C-12 fatty acids (2 × 10^-3M, pH 4.85). The inhibitory effect of the compound was dependent on the length of the carbon chain. Maximum effects were obtained for acids with eight to twelve carbon atoms per molecule. The inhibition was also dependent on the amount of undissociated acid present. By raising the pH so that the fatty acid dissociated the established inhibition was partly reversed. The effect of the neutral compound methyl octanoate was in essence identical to that obtained with octanoic acid. After fatty acid addition a close correspondence was found between the degree of inhibition of the oxygen uptake and the amount of UV absorbing substances leaking out from the cells. This extracellular material had an absorption maximum at 260 nm and a minimum around 240 nm. The leaking was ascribed to interaction between fatty acids or methyl octanoate and lipophilic substances of the cytoplasmic membrane. It is suggested that the inhibitory action on the endogenous respiration is due to similar effects on intracellular membrane systems.     

发酵法生产β-胡萝卜素菌丝体中氨基酸含量的分析

采用日立８３５－５０型氨基酸分析仪,测定了发醇法生产的β－胡萝卜素菌丝体及提取β－胡萝卜素后剩余的菌渣中氨基酸含量,结果表明：在氨基酸总含量中,必需氨基酸含量较高;在β－胡萝卜素菌丝体、菌渣中,赖氨酸含量较高,谷氨酸含量最高。     

Interaction of ascorbic acid and colchicine with sucrose utilization byCunninghamella elegans

The stimulating effect of ascorbic acid, administered to colchicine-treated mycelia ofCunninghamella elegans, on respiration, keto acid production and carbohydrate accumulation was lower than the highly increased rate of sugar absorption. Furthermore, the increasing ascorbic acid concentration in colchicine media favoured lower carbohydrate accumulation, keto acid production, carbon dioxide output and highly stimulated the utilization of the absorbed sugars for the formation of non-carbohydrate compounds. In addition, the vitamin seemed to counteract the inhibitory effects of colchicine on nucleoprotein and soluble nucleotide synthesis by colchicine-treated mycelia particularly when the drug was administered at high levels.     

Respiration of Bdellovibrio bacteriovorus Strain 109J and Its Energy Substrates for Intraperiplasmic Growth

Measurements of oxidation rates, respiratory quotients (RQ), and release of (14)CO(2) from uniformly labeled substrates showed that glutamate, alpha-ketoglutarate, and synthetic and natural amino acid mixtures are oxidized by suspensions of Bdellovibrio bacteriovorus strain 109J. The oxidation of these substrates largely suppress the endogenous respiration of the Bdellovibrio cells and may or may not cause a small increase, 20 to 50%, in their rate of oxygen consumption. The failure of respired substrates to increase markedly the respiration rate of the Bdellovibrio cells over the endogenous value is discussed. Carbon from these substrates is incorporated into the Bdellovibrio cells during oxidation. Acetate is also oxidized, but its oxidation inhibits endogenous respiration by only about 40% and no acetate is assimilated. The RQ of the Bdellovibrio cells changes from a value characteristic of endogenous respiration to that characteristic of the oxidation of glutamate or of a balanced amino mixture very shortly after the attack of the Bdellovibrio cells on their prey, and the latter RQ is maintained during intraperiplasmic growth. Glutamate, or a mixture of amino acids in the external environment, contributes to the carbon dioxide produced by the Bdellovibrio cells growing intraperiplasmically. It is concluded from these data that amino acids, derived from the breakdown of the protein of the prey, serve as a major energy source during intraperiplasmic growth of B. bacteriovorus 108J. Insofar as they were tested, B. bacteriovorus strains 109D and A. 3. 12 were similar in respiration to strain 109J.     

Differential impact of amino acids on OXPHOS system activity following carbohydrate starvation in Arabidopsis cell suspensions

Plant respiration mostly depends on the activity of glycolysis and the oxidation of organic acids in the tricarboxylic acid cycle to synthesize ATP. However, during stress situations plant cells also use amino acids as alternative substrates to donate electrons through the electron‐transfer flavoprotein (ETF)/ETF:ubiquinone oxidoreductase (ETF/ETFQO) complex to the mitochondrial electron transport chain (mETC). Given this, we investigated changes of the oxidative phosphorylation (OXPHOS) system in Arabidopsis thaliana cell culture under carbohydrate starvation supplied with a range of amino acids. Induction of isovaleryl‐CoA dehydrogenase (IVDH) activity was observed under carbohydrate starvation which was associated with increased amounts of IVDH protein detected by immunoblotting. Furthermore, activities of the protein complexes of the mETC were reduced under carbohydrate starvation. We also observed that OXPHOS system activity behavior is differently affected by different amino acids and that proteins associated with amino acids catabolism are upregulated in cells following carbohydrate starvation. Collectively, our results support the contention that ETF/ETFQO is an essential pathway to donate electrons to the mETC and that amino acids are alternative substrates to maintain respiration under carbohydrate starvation.     

猪苓营养菌丝与栽培菌核蛋白质成分的分析

对猪苓(Polyporus umbellate)营养菌丝与栽培菌核中蛋白质的氨基酸种类及含量进行了测定。结果表明,猪苓营养菌丝中各物质质量分数是:粗蛋白325.9g·kg-1,必需氨基酸总量34.924 g·kg-1,氨基酸总量91.956 g·kg-1;栽培菌核中各物质质量分数是:粗蛋白49.1 g·kg-1,必需氨基酸总量8.465 g·kg-1,氨基酸总量35.847 g·kg-1。营养菌丝中所含粗蛋白、氨基酸的量显著高于栽培菌核中粗蛋白、氨基酸的量。     

AMINO ACID DISTRIBUTION AND INCORPORATION INTO PROTEINS IN ISOLATED, ELECTRICALLY-STIMULATED CEREBRAL TISSUES

—The uptake of radioactive amino acid by incubated cerebral cortex slices is found to be a first order process. Incorporation of the radioactive amino acid into tissue protein is from a precursor pool that has first equilibrated with the intracellular endogenous free amino acids. Ways of calculating the amino acid incorporation in molar quantities from the observed incorporation of radioactivity are discussed, and it is concluded that the specific radioactivity of the intracellular acid-soluble fraction is the best basis for such estimates. The in vitro incorporation of leucine into tissue protein is estimated to be approximately 1±2 mμnol/mg protein/h, and of valine 0±4 mμmol/mg protein/h. Addition of free amino acids to the media had little or no effect on the calculated rates of incorporation. On incubation for 1 h the total free valine in tissue and medium increased by 0±43 μmol/g and leucine increased by 0±55 μmol/g. Estimates of amino acid incorporation based on the specific radioactivity of the media amino acids can give misleading results if this considerable release of amino acids into the medium is not taken into account. Electrical stimulation of neocortical slices with a variety of types of pulses was either without effect or decreased incorporation into portein. The decrease could not be directly correlated with changes in tissue K⁺ nor with the utilization of ATP. Mild, local stimulation of the lateral olfactory tract of piriform cortex slices was without effect on tissue phosphocreatine, K⁺ or amino acid incorporation.     

Entamoeba histolytica. I. Aerobic metabolism

The respiration of intact trophozoites harvested from axenic cultures of Entamoeba histolytica was studied with the polarographic technique utilizing the Clark oxygen electrode. A typical Q_o2 value for the freshly harvested amebae was 1 μatom oxygen/mg protein/hr.It was conclusively demonstrated that this parasite, a putative anaerobe, not only consumes oxygen when provided, but has a high affinity for the gas.Added glucose, galactose, and ethanol increased the respiratory rates, whereas other carbohydrates were without effect on the endogenous respiration. Intermediates of the tricarboxylic acid cycle, amino and fatty acids did not stimulate the respiration of E. histolytica.Inhibitors of the mammalian respiratory chain (cyanide, antimycin) as well as agents that inhibit enzymes catalyzing the tricarboxylic acid cycle (malonate, fluoropyruvate, fluoroacetate, fluorocitrate) had little effect on the endogenous or glucose-supported respiration. Alkylating agents (iodoacetamide, iodoacetate), cinnamate, and N-ethylymaleimide strongly inhibited the oxygen consumption of E. histolytica. The chemotherapeutic agents, Paromomycin, Emetine and Metronidazole, at concentrations that inhibit growth in vitro, did not restrict the respiration.Storage of the trophozoites at 4 C led to progressive deterioraion of the parasites and loss of endogenous and glucose-supported respiration. The deterioration was paralled by loss of SH-materials from the amebae. Likewise, sonication or lysis with detergents abolished both the endogenous respiration and response to glucose.Exogenous NADH or NADPH evoked only marginal increases in oxygen consumption of the freshly harvested amebae, but were effective respiratory substrates with stored or sonicated organisms. Addition of vitamin K₃ greatly enhanced the endogenous and glucose-supported respiration of the intact amebae, as well as enhancing the response of stored or sonicated amebae to reduced pyridine nucleotides.     

Quorum sensing activity in Ophiostoma ulmi: effects of fusel oils and branched chain amino acids on yeast-mycelial dimorphism

Aims: For Ophiostoma (Ceratocystis) ulmi, the ability to undergo morphological change is a crucial factor for its virulence. To gain an understanding of quorum‐sensing activity in O. ulmi as it relates to yeast‐mycelium dimorphism control, this study examines the effects of branched‐chain amino acids as well as their fusel alcohols and fusel acids as quorum sensing molecules. Methods and Results: In a defined medium containing glucose, proline and salts, O. ulmi grew as yeasts when the culture was inoculated with a high density of spores (2 × 10⁷ CFU ml^?1) and as mycelia when inoculated with a low spore density (4 × 10⁵ CFU ml^?1). The cultures displaying yeast morphology secreted a quorum‐sensing factor that shifted the morphology from mycelia to yeast. This quorum‐sensing molecule was lipophilic and extractable by organic solvents from the spent medium. Using GC/MS analysis, it was determined that the major compound in the extract was 2‐methyl‐1‐butanol. A similar effect was observed when the branched‐chain amino acids (fusel alcohol precursors) were used as the nitrogen source. E, E‐farnesol had no effect on the morphology of O. ulmi. Conclusions: Addition of the branched‐chain amino acids or one of the compounds detected in the spent medium, 2‐methyl‐1‐butanol or 4‐hydroxyphenylacetic acid, or methylvaleric acid, decreased germ tube formation by more than 50%, thus demonstrating a quorum sensing molecule behaviour in O. ulmi cultures. Significance and impact of the study: This study presents advances in the investigation of dimorphism in O. ulmi, complementing the existing scientific basis, for studying, understanding and controlling this phenomenon.     

Regulation of formation of aerial mycelia and spores of Streptomyces viridochromogenes.

Growth of Streptomyces viridochromogenes on a solid glycerol-NH4NO3 salts medium was accompanied by the formation of aerial mycelia and spores. Adding 0.5% or more casein hydrolysate to the medium stimulated growth while completely repressing the formation of aerial mycelia and spores. This repression was temporary, as evidenced by the fact that transfer of the organisms to media not containing casein hydrolysate resulted in the appearance of aerial mycelia and spores. The effects of individual amino acids were tested. Glycine retarded growth and repressed formation of both aerial mycelia and spores. L-Aspartic acid, L-glutamic acid, and L-histidine stimulated or had little effect on growth and repressed formation of spores but not aerial mycelia. Repression by casein hydrolysate could not be attributed to the carbon/nitrogen ratio or the pH of the medium. Adding 1.25 to 2.5 mM adenine to the medium caused a reversal of the casein hydrolysate repression of aerial mycelium formation but did not reverse repression of sporulation. Dimethyladenine and 8-azaguanine had an effect similar to that of adenine, but a variety of other purine or pyrimidine derivatives had no effect on casein hydrolysate repression. The repression of aerial mycelium and spore formation by casein hydrolysate occurred only in media containing 15 mM or more phosphate. Aerial mycelia and spores were formed in media containing casein hydrolysate and 3 mM or less phosphate.     

樟芝菌丝体的氨基酸成分分析

测定了樟芝发酵菌丝体中游离氨基酸和结合氨基酸的含量。结果表明,樟芝菌丝体中含有丰富的8种人体必需氨基酸、支链氨基酸和谷氨酸,其中8种必需氨基酸含量是FAO(联合国粮农组织)标准的5.55倍,而芳香族氨基酸的含量却很低。在测定过程中,由于采用了酸水解,菌丝体中的色氨酸被破坏,而其余17种氨基酸均被测出,含量极其丰富。尤其是支链氨基酸的含量远大于α-酪蛋白、卵蛋白和大豆球蛋白。