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Distribution of enzymes in cell-free yeast extracts

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... The failure to prepare cell-free, actively respiring yeast extracts contributes to the continuing controversy about the main respiratory mechanism of this micro-organism . . . ([120] p. 62). Eventually, however, respiring mitochondria were prepared from baker's yeast. ...
Article
The cell knows two methods of getting out the energy of [foodstuff] molecules; it either fragments them or burns them. The first method we refer to as fermentation, the second, oxidation.
... The method of grinding was one of the chief difficulties in this study. The grinding must be vigorous in order to break the tough cell walls that internal structures may also be disrupted, a problem which has been emphasized by others (7,23). Also, the incomplete cell breakage makes it difficult to study the coarse dCbris fraction, especially since alumina is also present. ...
Article
The cellular distribution of pyruvate decarboxylase and acetyl-CoA kinase in C. pulcherrima grown on glucose has been investigated. By using a mild procedure for the separation of the cytoplasmic and mitochondrial fractions, it could be demonstrated that both enzymes are almost exclusively localized in the cytoplasm. The levels of pyruvate decarboxylase in Candida pulcherrima and Saccharomyces cheresiensis grown aerobically on different carbon sources have also been studied: it was high in cells from glucose, glucose plus acetate, or glucose plus pyruvate, and low in cells from acetate or pyruvate. By contrast, the content of acetyl-CoA kinase was always relatively constant. Evidence is also presented for the induction of pyruvate decarboxylase by glucose.
Article
1.1. A procedure for the isolation of actively respiring particles from yeast is described. These particles are considered to be mitochondria. They stain vitally with Janus green B and oxidize the main citric acid cycle substrates as well as pyruvate and acetate.2.2. The washed mitochondria oxidize pyruvate only in the presence of a catalytic amount of one of the Krebs cycle substrates. In the presence of catalytic amounts of malate, pyruvate is oxidized to carbon dioxide and water.3.3. The requirements for acetate oxidation are more specific than those for pyruvate. Unlike pyruvate, only α-ketoglutarate and citrate initiate acetate oxidation. The pathway by which acetate is dissimilated has not been established.
Article
1.1. A high-speed shaker has been used for the preparation of actively respiring extracts of baker's yeast, Aerobacter aerogenes and Proteus vulgaris OX 19. Disintegration periods ranged from 10 to 40 seconds.2.2. 10-second yeast extracts contain granules which oxidise isocitrate, α-ketoglutarate, succinate, lactate and ethanol, and which account for a large proportion of the respiration of the whole extract.3.3. 15-second extracts of Proteus oxidise many substrates, and the isolated granules oxidise succinate, lactate and formate. With lactate as substrate, whole extracts and, to a lesser extent, high-speed supernatants, show a phosphorylation sensitive to 2,4-dinitrophenol; the granules alone show no phosphorylation.4.4. 15-second extracts of Aerobacter also oxidise a variety of substrates, but the isolated granules oxidise succinate and lactate only. Whole extracts and high-speed supernatant fractions, but not the granules, show a phosphorylation insensitive to 2,4-dinitrophenol with glucose as the substrate. The granules have slight phosphorylating activity with lactate as substrate.5.5. The relation of the granules of micro-organisms to animal mitochondria is discussed.
Article
Yeast cell fractions prepared in buffer show a random distribution of lipide-synthesizing enzymes. When the cell fractions are obtained with the aid of a saturated lactose solution to afford protective conditions, most of these enzymes are found in the particulate, or mitochondrial, fraction which is sedimentable by a centrifugal force of 11,500 × g.A requirement of coenzyme A (CoA) for sterol synthesis from acetate by cell-free yeast extracts can be demonstrated only when the extracts are partially depleted of the cofactor by treatment with Dowex 1-Cl. Further supplementation with CoA is required for the conversion of squalene to the fatty acids.
Article
1.1. Respiring cell-free baker's yeast extracts have been prepared by 10-sec high-speed mechanical disintegration.2.2. Increasing the disintegration period reduces the oxidate ability of the extracts.3.3. Whole extracts have a low blank respiration at or above pH 7·4. At pH 6·5–7·1 a high blank respiration is found. Addition of small amounts of ATP or A–5–P to the blank at pH 7·4 also induces this high respiration.4.4. The blank respiration is due to catabolism of polysaccharide. The R.Q. of the reaction is about 1 or sometimes higher. Considerable amounts of pyruvate and acetate are formed, but these are insufficient to account for either carbohydrate disappearance, O2 uptake, or CO2 formation. Some pentose formation occurs.5.5. Various acids of the Krebs tricarboxylic acid cycle are oxidized by the extracts, succinate most and malate least rapidly.6.6. Quantitative balances of the metabolism of each of the Krebs cycle acids show that they can largely be accounted for by the reactions of the cycle. Only small amounts of citrate are formed from oxaloacetate, acetate and CoA. Considerable amounts of glutamate are formed from α-ketoglutarate but not from citrate.7.7. The results are discussed in the light of the Krebs cycle being a major pathway of yeast respiration. Other pathways exist and may account for the high blank respiration observed under certain conditions.
Article
The rates of release of 7 enzymes from bakers' yeast have been measured. The disruption process did not cause loss of activity of these enzymes. The various operating pressures, temperatures, and initial yeast concentrations used did not affect the rates of enzyme release relative to protein release. The release of acid phosphatase and invertase was faster than the overall protein release. Alcohol, glucose-6-phosphate, and 6-phosphogluconate dehydrogenases were released slightly faster or at the same rate as the overall protein and alkaline phosphatase and fumarase were released more slowly. These observations correlate well with the reported locations of these enzymes in the yeast cell.
A specially designed high-speed blendor and glass beads have been used to disintegrate yeast cells. The method enables large quantities of cells to be fragmented quickly at low temperature, and cell-free mitochondrial particles to be prepared in high yield. The particles are isolated in a sucrose-Tris-EDTA medium and extensively refractionated in the same medium. The success of the fractionation is dependent upon the presence of the Tris buffer, as the latter prevents the aggregation of the particulate material. Two morphologically and enzymatically different particle types have been obtained: a heavy fraction corresponding to mitochondria in size and internal organization, and a light fraction consisting of vesicular, single-membrane particles of a smaller size. The light particles oxidize DPNH and succinate, but do not oxidize pyruvate-malate, and lack the capacity for phosphorylation. The heavy particles oxidize pyruvate-malate as well as the citric acid cycle intermediates, although their alpha-ketoglutaric dehydrogenase activity is low. Oxidation by the heavy particles is coupled to phosphorylation, and P/O ratios of about 1.5 have been obtained. Lactic acid dehydrogenase is also present in the heavy fraction, and lactate is oxidized with a P/O ratio of about 0.7.
Article
Mitochondrial and cytoplasmic fractions of baker's yeast were prepared by mechanical disintegration from cultures grown rather anaerobically and in vigorously aerated conditions. The mitochondrial fraction contained only 25–26% of the succinic acid dehydrogenase activity and the succinic acid oxidase activity.The total thiamine content of the mitochondria (including the disrupted ones) amounted to 6.0 μg for the anaerobic cultures and for the aerobic cultures to 5.1 μg per g fresh yeast. The total content of thiamine in the cytoplasm was found to be 1.5 μg at the anaerobic stage and a scant 0.3 μg per g fresh yeast at the aerobic stage.80–95 % of the thiamine is located in the mitochondria. During transfer from anaerobic to aerobic culture conditions, the mitochondrial content of thiamine decreases by a scant 20%, whereas the cytoplasmic content falls by a good 80%. This sharp decrease in the thiamine content of the cytoplasm is consistent with the earlier observation that the decarboxylase activity of baker's yeast strongly decreases on transference from anaerobic to aerobic culture conditions.
Article
Protoplasts of Lipomyces lipofer were ruptured by decompression in a French pressure cell. A particulate fraction sedimenting at 17,000 × g in saline or sucrose media contained the bulk of the substrate-dependent oxidative activity and was capable of phosphorylation. Particle fractions isolated in sucrose required supplementation with ATP and Mg2+ while fractions from saline isolations required, in addition, NAD and cytochrome c. NADP and thiamine pyrophosphate had small or negligible effects in the presence of the other cofactors. Oxidative phosphorylation occurred most efficiently in particles isolated in sucrose, but in no case did the P/O ratio exceed 1.6. Electron microscopic examination of the sucrose-isolated fraction showed it to be a relatively homogeneous preparation of mitochondria which appear more "native" after incubation with substrate than immediately after isolation.
Article
1. 1. Some changes in the structure and metabolism of yeast cells which occur on freezing with solid CO2 are described. 2. 2. CO2-frozen cells, besides being permeable to addrd organic acids, lose organic acids, amino acids, protein, carbohydrate, coenzymes and inorganic salts when washed with water or buffer. 3. 3. The ability of such cells to respire ethanol and succinate may not be a reliable guide to the normal metabolism of these substrates, since the response to varying degrees of temperature-shock differs according to the strain of bakers' yeast used. 4. 4. Freezing reduces the amount of sedimentable material in cell-free extracts prepared by ultra-rapid shaking. In this respect, the effects of freezing resemble those obtained by disintegrating fresh cells for longer periods. Partial solubilization of granular fumarase, aconitase, and ethanol and malic dehydrogenases occurs, and aconitase is partially inactivated. 5. 5. The coenzyme-independent ethanol and malic dehydrogenase activities found in extracts of fresh yeast are lacking in extracts of frozen yeast. The possibility of coenzyme-binding, the disruptive action of thermal shock and the evidence for the participation of the tricarboxylic acid cycle in acetate oxidation by bakers' yeast, are discussed.
Article
The isolation, purification, physical and chemical properties of a mitochondrial DNA of Saccharomyces cerevisiae are described and compared with the nuclear DNA of the same organism. The two entities are clearly distinct in absorption spectra at various pH values, thermal transition temperature in two different solvent systems, buoyant density in CsCl and base composition, all of which are consistent with a GC content of 35 % for nuclear and 21 % for mitochondrial DNA. The latter, a normal double helical molecule, occurs integrated into respiratory particles which renders it DNase resistant. It is present to the extent of 0.71 μg per mg of particle protein in highly aerobic cells of the wild type, decreases during glucose repression and is greatly reduced (⩽ 0.06 μg/mg protein) in a cytoplasmic respiratory-deficient mutant (vegetative petite, ϱ−). The So20,w of the isolated DNA equals 33 s, hence its molecular weight is approximately 2 × 107. Evidence is also presented for its preferential interaction with acridines. The implications of all these findings in the problems of mitochondrial autonomy and self-duplication are discussed.
Article
Mitochondrial particles, to which the bulk of the succinoxidase and cytochrome oxidase activities were exclusively bound, were isolated from Candida albicans after disrupting the growing cells in a mortar with quartz sand. Electron-microscopic examination of the mitochondrial preparation showed that it mainly contained native and submitochondrial particles. Mitochondria thus prepared could couple phosphorylation to oxidation of various substrates including TCA cycle members, exogenous NADH, glutamate, ethanol, and D- and L-lactate. P/O ratios obtained with these substrates corresponded closely to those reported for mitochondria from other fungi, approximately 1 unit lower than P/O ratios with mammalian intact mitochondria, whereas much lower respiratory control ratios were observed. Oligomycin strongly inhibited oxidative phosphorylation and Mg++-induced ATPase activity. The effects of inhibitors of electron transfer such as amytal, rotenone, antimycin A or sodium azide on Candida mitochondria were similar to those on mammalian mitochondria and it is of note that the sensitivity of oxidation of both a-ketoglutarate and NADH by C. albicans mitochondria to rotenone exceeded those of any mitochondrial preparations of other fungi.
Article
Cells of Saccharomyces carlsbergensis H 60 synthesize alcohol dehydrogenase (ADH) in media containing 2% lactat, 1% ethanol or 0.1% glucose. Crystals may be induced in protoplasts of these cells. Increase of glucose concentration in the medium results in diminished ADH synthesis and decreased tendency for crystal formation. Repression of ADH synthesis by glucose results in the formation of a protein (MG 110000 D), the significance of which is discussed. Early stages of crystal formation inside the cell are demonstrated electronmicroscopically. At first dense material accumulates between opposite membranes of neighbouring mitochondria. Within mitochondria frequently membrane bundles occur in close vicinity to crystals. These ADH‐crystals arise from this material.
Chapter
The enzyme which catalyses the equilibrium expressed by the reaction was discovered by Martius and Knoop 1 and named aconitase by Breusch 2. It has been suggested that two enzymes are concerned in catalysing this reaction3–8, but although aconitase has not been isolated in the pure state, the available evidence indicates that only a single enzyme is involved9–16. Recently, it has been established that of the four stereoisomers of isocitric acid only the dextrorotatory L s isomers occurs naturally17. (L s corresponds to the configuration of the a carbon of naturally occurring serine.)
Chapter
Energy required for vital function whether in the animal, the bacterium or the higher plant is obtained by burning fuel. In most plant tissues the fuel is sugar or one of its storage products (either di- or polysaccharides). In other instances it is fat, and in a few plants, protein storage products may be burned to provide energy. The energy available in these molecules, in the form of bond energy, is derived from the sun either directly or indirectly. For this energy to become available to the cell, a series of transformations must occur which, in the case of the sugars and their related products, and for at least part of the fat molecule, involves the formation of phosphorylated compounds. In short, if substrates are to be used as energy sources they must first undergo phosphorylation. This appears to be true for almost all instances of sugar oxidation. There are, however, a few isolated cases in some animal tissues and especially in bacteria where prior phosphorylation may not be required.
Chapter
This chapter discusses two ways of releasing mitochondria from within the yeast cell walls (1) break the cell wall open by force; (2) remove the cell wall by enzymatic digestion. The various methods available and their relative merits are described in the chapter. The most frequently used methods for breaking cell walls by force are those employing small glass beads with either rapid stirring or shaking. Mitochondria are also prepared from yeast cells by either ultrasonic treatment, grinding with Carborundum, or by the use of a French press. An improved method of high-pressure extrusion, which employs the Sorvall Ribi Cell Fractionator, is also used successfully. This instrument allows large quantities of yeast cells to be processed, and the temperature at the point of breaking is carefully controlled by a stream of refrigerated nitrogen. Enzymes from the gut juice of Helix pomatia are used for the digestion of yeast cell walls. The mitochondrial fraction prepared is further purified by density gradient centrifugation. The different methods of preparation described produce mitochondria with varying degrees of structural integrity and differing properties. Yeast mitochondria purified by density gradient flotation contain DNA. A DNA-dependent synthesis of RNA has been described in yeast mitochondria.
Article
One of the major recent advances in cellular architectonics, the spatial and structural organization of reactions and processes, is the elucidation of the role of mitochondria in cell metabolism. Plant cell is not simply a bag of haphazardly arranged enzymes. The cell particulates possess a high degree of structural and functional organization, which is, under certain conditions, very labile. The fundamental processes carried out by the mitochondria appear to be the terminal transfer of electrons, the coupling of energy-trapping mechanisms (phosphorylations) to oxidations, and the Krebs cycle reactions and the numerous ancillary processes that eventually funnel through the cycle. In spite of the remarkable autonomy of the mitochondria, the overall activities of the cell are the result of intimate interactions among the various cellular components. Glycolysis involves the plastids, the soluble fraction, and possibly the nucleus as well as the mitochondria1 enzymes. Certain enzymes involved in the Krebs cycle (e.g., malic dehydrogenase) and in hydrogen transfer (e.g., cytochrome reductase) are not confined entirely to the mitochondria. The mitochondria interact with the nucleus in phosphorylation, with the chloroplasts in photosynthesis, and with the microsomes in protein synthesis. In close relation to the outer cell boundary, the mitochondria may participate actively in the movement of substances into the cell or in the growth of the cell wall.
Article
This chapter focuses on the recent approaches to the cytochemical study of mammalian tissues. The methods of correlation of cytological and biochemical data are discussed. The cell fractionation technique depends on the cytological identification of the structural components present in the cell fractions and the assignment of specific biochemical properties to each of the intracellular elements that can be identified cytologically within the intact cell. Two important and new techniques are employed with some success in approaching the problem of the cytological identification of cytoplasmic components. The first is examination of the isolated structures in the electron microscope after appropriate fixation, embedding, and the cutting of ultrathin sections. The second involves the use of new methods of preparative centrifugation by which the resolving power in the separation of particles of differing size and density has been enormously increased. These technical advances pointed up the need for, and provided a means of approach to, the development of media that permits the intracellular structures to retain their normal morphological characteristics upon disruption of the cell, and at the same time permits their isolation in homogeneous preparations.
Chapter
This chapter focuses on the preparation and solubilization of bacterial particles. In contrast to the activity found with crude bacterial homogenates or mammalian mitochondria, the isolated particles from bacteria usually exhibit an additional requirement for factors which were apparently solubilized during the disruptive procedure. Bacterial systems that can be fractionated and reconstituted by the addition of essential components liberated from complex structures offer new tools for elucidating the mechanisms involved in oxidative phosphorylation and in protein biosynthesis. Although a large number of enzymes are associated with the large particulate fraction, the preparations are assayed only for their ability to couple phosphorylation to oxidation, since this activity is labile and can be used as an index of structural integrity. The particulate and supernatant fractions are separated and then tested for their ability to couple phosphorylation to oxidation. The assay system consists of the bacterial fractions, either separately or reeombined, a suitable electron donor, orthophosphate, and a phosphate aceeptor system. The consumption of oxygen and the esterification of orthophosphate are measured.
Article
A personal historical view of the biochemistry of glucose catabolism in yeast and muscle by Dr J.A. Barnett.
Aerobic yeast mitochondria contain a unique isozyme of alcohol dehydrogenase that accounts for the ability of yeast mitochondria to respire with ethanol as a substrate. This alcohol dehydrogenase has been isolated from intact yeast mitochondria, purified approximately 65-fold, and further studied. The mitochondrial isozyme represents about 5% of the total cellular alcohol dehydrogenase and differs from the classical, cytoplasmic isozyme by its slower electrophoretic mobility on polyacrylamide gel and its more alkaline pH optimum. Only minor differences between the mitochondrial and cytoplasmic isozymes were noted in the apparent for NAD+, NADH, ethanol, and acetaldehyde.
Article
1.1. The presence of a pyruvic oxidase in yeast has been demonstrated. This system is similar to that described for Escherichia coli and Streptococcus faecalis.2.2. This system is present both in the soluble and in the granular fraction. It is also present in yeast after anaerobic growth and in a mutant produced by respiratory deficiency.3.3. The action of ethanol, acetaldehyde, arsenous ions, and a dimercaptol, has been studied.4.4. The significance of this system is discussed.
Article
Cells of Saccharomyces carlsbergensis H 60 synthesize alcohol dehydrogenase (ADH) in media containing 2% lactat, 1% ethanol or 0.1% glucose. Crystals may be induced in protoplasts of these cells. Increase of glucose concentration in the medium results in diminished ADH synthesis and decreased tendency for crystal formation. Repression of ADH synthesis by glucose results in the formation of a protein (MG 110000 D), the significance of which is discussed. Early stages of crystal formation inside the cell are demonstrated electronmicroscopically. At first dense material accumulates between opposite membranes of neighbouring mitochondria. Within mitochondria frequently membrane bundles occur in close vicinity to crystals. These ADH-crystals arise from this material.
Article
A specially designed high-speed blendor and glass beads have been used to disintegrate yeast cells. The method enables large quantities of cells to be fragmented quickly at low temperature, and cell-free mitochondrial particles to be prepared in high yield. The particles are isolated in a sucrose-Tris-EDTA medium and extensively refractionated in the same medium. The success of the fractionation is dependent upon the presence of the Tris buffer, as the latter prevents the aggregation of the particulate material. Two morphologically and enzymatically different particle types have been obtained: a heavy fraction corresponding to mitochondria in size and internal organization, and a light fraction consisting of vesicular, single-membrane particles of a smaller size. The light particles oxidize DPNH and succinate, but do not oxidize pyruvate-malate, and lack the capacity for phosphorylation. The heavy particles oxidize pyruvate-malate as well as the citric acid cycle intermediates, although their α-ketoglutaric dehydrogenase activity is low. Oxidation by the heavy particles is coupled to phosphorylation, and P/O ratios of about 1.5 have been obtained. Lactic acid dehydrogenase is also present in the heavy fraction, and lactate is oxidized with a P/O ratio of about 0.7.
A simple procedure for the disintegration of yeast cells, by which litre quantities of cell extract (approximately 11 per cent dry weight) may be obtained, is described. The fresh yeast cake is treated with dry ice in a high speed electric homogenizer after which it is left to thaw out at 0°. No addition of buffer has to be made, and the cell juice obtained after centrifugation of the resulting slurry appears to contain different subcellular particles and various enzymes in a relatively native state.When the extract is incubated at higher temperatures, proteolysis will rapidly change the electrophoretic and enzymatic properties, which indicates that procedures involving autolysis may be dangerous when information about the chemical composition of the native enzymes is desired.
Chapter
IntroductionLe cycle tricarboxylique chez les microorganismeLes biosynthèses cellulaires liées au cycle des acides tricarboxyliquesLe cycle des acides tricarboxyliques en anaérobioseConclusions
Article
In order to evaluate the fundamental behaviour of the fungi Neurospora crassa, Oospora lactis and Saccharomyces cerevisiae in the cytochemical detection of dehydrogenases, detailed experiments were carried out concerning the choice of the most suitable tetrazolium salt, the influence of N,N-dimethylformamid as solvent for the indicators, the value of the addition of intermediators (phenazine methosulfate, menadione) and of auxiliary substances (MgCl2, KCN, NaN3, amytal, polyvinylpyrrolidone) to the incubation medium and the importance of the “nothing dehydrogenase” activity. The importance of control reactions and the resulting observations were also investigated. The results of these basic experiments were summarized in a list of incubation media for the detection of specific dehydrogenases. The tetrazolium salts Nitro-BT and Tetranitro-BT proved to be the best indicators. The possibility of an exact intracellular enzyme localization in the cells of the three fungi are discussed.
Article
(1) Intact cells of baker's yeast are impermeable to succinate and ferricyanide so that their suspension does not show any succinic dehydrogenase activity with ferricyanide as electron acceptor. (2) On incubation with sodium deoxycholate the cells become permeable and their suspension shows succinic dehydrogenase activity. (3) The effect of sodium deoxycholate on the change in yeast permeability is markedly dependent on temperature, pointing to a high temperature coefficient. The effect is higher at lower pH values (unless free deoxycholic acid is precipitated from the solution) and at higher ionic strength. (4) Sodium deoxycholate concentrations below 0.002 M are completely ineffective under the given experimental conditions, at 0.005 M the effect is already maximal. (5) Cholate is much less effective than deoxycholate; dehydrocholate was completely ineffective under the experimental conditions used. (6) The results indicate that the high effectiveness of deoxycholate is related to its ability to form polymeric helical complexes.
Article
1. Mitochondria exhibiting respiratory control were prepared from bakers' yeast and from Saccharomyces cerevisiae strain D-261 after disrupting the cells in a colloid mill at low speeds with glass beads. Optimal values of the variables involved were determined. Yields of 13–25 mg mitochondrial protein per 100 g packed cells were routinely obtained. Mitochondrial quality was similar to that of particles prepared by a method employing snail gut enzymes to digest cell walls.2. The functions of mitochondria prepared under the mildest homogenization conditions were only slightly affected by large alterations in pH or Mg2+ concentration of the reaction medium.3. Oligomycin and atractyloside were potent inhibitors of phosphorylating respiration in yeast mitochondria.4. Respiration was specifically stimulated by ADP, with an apparent Km = 26.5 μM. Phosphate also stimulated malate-supported respiration, with an apparent Km of 1.5 mM. Varying the phosphate concentration with succinate as substrate led to a transition from ADP-controlled respiration at high phosphate to uncontrolled respiration at low phosphate.5. P:O or ADP:O values for this preparation were about 1.8 during succinate oxidation, 1.8 during ethanol oxidation, 2.4 during α-ketoglutarate oxidation and 1.7 during oxidation of malate plus pyruvate.
Article
1. The activities of the enzymes of the citric acid cycle, the glyoxylate by-pass and some other enzymes acting on the substrates of these cycles have been measured at the pH of the yeast cell during the aerobic growth of yeast on different carbon sources and in different growth media. 2. Sugars induced an anaerobic type of metabolism as measured by ethanol production. Glucose was much more effective in inducing the anaerobic pathways than was galactose. The production of ethanol by cells grown on pyruvate was very small. 3. Glucose was also a more effective repressor than was galactose of the citric acid-cycle enzymes but both were equally effective in repressing almost completely the enzymes of the glyoxylate by-pass. 4. Disappearance of the sugars from the growth medium resulted in an increase in the activities of the enzymes of the citric acid cycle and in the appearance of substantial activities of the enzymes of the glyoxylate cycle. By contrast, the activities of purely biosynthetic enzymes (glutamate-oxaloacetate transaminase, NADP(+)-linked glutamate dehydrogenase) and of pyruvate decarboxylase were decreased. 5. The 2-oxoglutarate-oxidase system was found to be the least active enzyme of the citric acid cycle. 6. The regulatory control at the levels of pyruvate and acetaldehyde and the control of the citric acid cycle are discussed.
Article
A comparative study of three enzymes, aconitase, fumarase, and DPN-linked isocitric dehydrogenase was made in normal baker's yeast and in the mutant "petite colonie" grown under varying conditions of oxygen supply. When both normal and mutant yeasts are grown anaerobically, aconitase, fumarase, and DPN-isocitric dehydrogenase activities in the two strains are alike. When grown in the presence of air, all three enzymes show increased activity, but under such conditions distinct differences between the two strains become apparent: the increases in the normal strain are much more pronounced than in the mutant strain. Aeration of non-proliferating suspensions of anaerobically grown normal yeast which is known to re-establish ability to respire also leads to an increased formation of aconitase and, to a lesser extent, fumarase, but in order to obtain their full complement of these enzymes, extensive growth of the cellsi n the presence of air seems necessary. By differential centrifugation all three enzymes were found to be associated mainly with the supernatant fraction in both normal and mutant yeast. The function of these enzymes in the metabolism of yeast is discussed.
Article
THE mitochondrial membrane has been a subject of controversy. Two distinct problems have arisen: whether a morphological membrane exists, and, if so, whether it is semi-permeable.
Article
A spectrophotometric method of measuring the enzymatic formation and disappearance of umaric and cis-aconitic acids is reported.RésuméNous décrivons une methode spectrophotométrique qui permet de mesurer la formation et la disparation enzymatique de l'acide fumarique et de l'acide cis-aconitique.ZusammenfassungEine spektrophotometrische Methode zur Messung der enzymatischen Bildung und Zerstörung von Fumarsäure und cis-Akonitsäure wird beschrieben.
Article
The experiments reported in this paper concern the mechanisms involved in plant respiration, particularly those involved in the oxidation of pyruvate. The interest of such a study lies in the fact that although pyruvate is well established as an intermediate in the respiratory oxidation of hexose by plant tissues,(1) it has not heretofore been possible to bring about the further oxidation of pyruvate in vitro by any enzyme system of plant origin.
  • O Warburg
  • W Christian
  • A Griese
Warburg, O., Christian, W. & Griese, A. (1935). Biochem. Z. 282, 157.
The Yeast Cell, its Genetics and Cytology
  • C C Lindegren
Lindegren, C. C. (1949). The Yeast Cell, its Genetics and Cytology. St Louis: Educational Publishers.
  • H M Hirsch
Hirsch, H. M. (1952). Biochim. biophys. Acta, 9, 674.
  • P M E Nossal
Nossal, P. M. (1953b). Biochim. biophys. Acta, 11, 596. Racker, E. (1950). Biochim. biophys. Ada, 4, 211.
  • J L Still
  • E H Kaplan
Still, J. L. & Kaplan, E. H. (1950). Exp. Cell. Res. 1, 403.
  • F Huenneckens
  • D E Green
Huenneckens, F. & Green, D. E. (1950). Arch. Biochem. 27, 416.
  • S Weinhouse
  • R H Millington
Weinhouse, S. & Millington, R. H. (1947). J. Amer. chem. Soc. 69, 3089.
  • S P Colowick
  • N Kaplan
Colowick, S. P., Kaplan, N. 0. & Ciotti, M. M. (1951). J. biol. Chem. 191, 447.
  • A H Kornberg
Kornberg, A. H. (1950). J. biol. Chem. 182, 805.