Terminal Oxidase System Research Articles

The physiology and metabolism of Campylobacter jejuni and Helicobacter pylori.

Although closely related phylogenetically and sharing many phenotypic similarities, Campylobacter jejuni and Helicobacter pylori are distinct in some major aspects of their physiology and metabolism. In this paper, comparative aspects of the physiology of these pathogens is discussed in the light of recent biochemical and genome sequence data. Campylobacter jejuni is emerging as a more versatile and metabolically active pathogen, with a complete citric acid cycle, and a complex and highly branched respiratory chain which allows both aerobic and anaerobic respiration with a variety of alternative electron acceptors. These properties enable it to survive in a number of environments in addition to the mammalian or avian gut. In vivo, anaerobic growth of C. jejuni could be an important factor allowing intestinal colonization. Helicobacter pylori is a more specialized pathogen, largely restricted to the human stomach, with a unique combination of virulence factors, an incomplete citric acid cycle, a simpler respiratory chain with only a single terminal oxidase and fewer regulatory systems. Both bacteria are microaerophiles and, while there is likely to be no single explanation for their oxygen sensitivity, there is evidence that the possession of oxygen-sensitive enzymes and an increased sensitivity to oxidative stress play an important role.

Respiratory control determines respiration and nitrogenase activity of Rhizobium leguminosarum bacteroids.

The relationship between the O2 input rate into a suspension of Rhizobium leguminosarum bacteroids, the cellular ATP and ADP pools, and the whole-cell nitrogenase activity during L-malate oxidation has been studied. It was observed that inhibition of nitrogenase by excess O2 coincided with an increase of the cellular ATP/ADP ratio. When under this condition the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) was added, the cellular ATP/ADP ratio was lowered while nitrogenase regained activity. To explain these observations, the effects of nitrogenase activity and CCCP on the O2 consumption rate of R. leguminosarum bacteroids were determined. From 100 to 5 microM O2, a decline in the O2 consumption rate was observed to 50 to 70% of the maximal O2 consumption rate. A determination of the redox state of the cytochromes during an O2 consumption experiment indicated that at O2 concentrations above 5 microM, electron transport to the cytochromes was rate-limiting oxidation and not the reaction of reduced cytochromes with oxygen. The kinetic properties of the respiratory chain were determined from the deoxygenation of oxyglobins. In intact cells the maximal deoxygenation activity was stimulated by nitrogenase activity or CCCP. In isolated cytoplasmic membranes NADH oxidation was inhibited by respiratory control. The dehydrogenase activities of the respiratory chain were rate-limiting oxidation at O2 concentrations (if >300 nM. Below 300 nM the terminal oxidase system followed Michaelis-Menten kinetics (Km of 45 +/- 8 nM). We conclude that (i) respiration in R. leguminosarum bacteroids takes place via a respiratory chain terminating at a high-affinity oxidase system, (ii) the activity of the respiratory chain is inhibited by the proton motive force, and (iii) ATP hydrolysis by nitrogenase can partly relieve the inhibition of respiration by the proton motive force and thus stimulate respiration at nanomolar concentrations of O2.

The unique steroidogenesis of the aldosteronoma in the differential diagnosis of primary aldosteronism.

18-Hydroxycortisol and 18-oxocortisol have been isolated from the urine of patients with aldosterone producing adrenocortical adenomas, but not from those with idiopathic hyperaldosteronism associated with bilateral adrenal hyperplasia. These C-18 oxygenated cortisols are biosynthesized by the substitution of cortisol for the normal substrate, corticosterone, in the terminal oxidase system required for the biosynthesis of 18-hydroxycorticosterone and aldosterone. To make use of this biochemical difference between the two groups in the preoperative diagnosis of primary aldosteronism, we have developed and utilized a specific primary standard analytical method, stable isotope dilution mass fragmentography, for quantifying 18-hydroxycortisol and the tetrahydro metabolite of 18-oxocortisol in 24-h urine samples. The normal range by this technique of 4.6 +/- 1.8 micrograms/day tetrahydro 18-oxocortisol and 43 +/- 23 micrograms/day 18-hydroxycortisol in urine was lower and narrower than previous estimates using other methods. Excretion of the 18-oxocortisol metabolite ranged from 2-12 micrograms/day in bilateral hyperplasia and 17-1203 micrograms/day in typical adenomas. 18-Hydroxycortisol excretion similarly separated bilateral hyperplasia (23-59 micrograms/day) from typical adenomas (60-2750 micrograms/day). The cortisol C-18 oxidation pathway describes a unique steroidogenic mechanism in the aldosteronoma not present in idiopathic aldosteronism due to bilateral adrenal hyperplasia and as such provides a basis for the biochemical classification of primary aldosteronism and the differentiation of these two groups. This unique biochemistry was also observed in unilateral hyperplasia but not in the renin-dependent aldosteronoma.

Bacteroids from soybean root nodules: respiration and N 2 -fixation in flow-chamber reactions with oxyleghaemoglobin

Suspensions of bacteroids prepared from root nodules of glasshouse-grown soybeans were studied in a stirred chamber supplied with variable flows of solutions containing dissolved air, oxyleghaemoglobin and various energy sources. In experiments of up to 6 h duration, several steady states were established in which frequent measurements were made of the concentration of free, dissolved O 2 in the range 5–200 nM and of rates of O 2 consumption, CO 2 efflux and N 2 fixation. The principal findings were: (i) bacteroids were capable of efficient N 2 fixation without the supply of exogenous energy sources when respiring at 10–50 nM free O 2 . This endogenous respiration was enhanced after periods of supply of succinate or malate. (ii) Exogenous energy sources were of three types, those that were poorly utilized (glucose), those that enhanced endogenous respiration and supported efficient N 2 fixation (glutamate, 2-oxoglutarate) and succinate and malate, which promoted respiration but supported N 2 fixation inefficiently or in some circumstances inhibited it. It is proposed that succinate and malate act primarily to increase endogenous reserves that become the principal source of reducing power for N 2 fixation, (iii) Bacteroids have a terminal oxidase system with very high apparent affinity for O 2 ( s 0.5 ≈ 5–8 nM) and complex kinetics (plots of v against s are sigmoidal; n app > 1.8). This system was active with endogenous substrates and when glutamate or 2-oxoglutarate were supplied. With succinate or malate, respiration appeared to be the sum of endogenous activity plus O 2 consumption by a system with lower affinity for O 2 ( K s = 38 nM) and simple kinetics ( n app ≈ 1). (iv) During the first hour of reactions there were changes in O 2 demand and oscillations in O 2 demand and CO 2 efflux followed supply or withdrawal of exogenous substrates. It is proposed that these changes are examples of phenotypic plasticity in these symbiotic bacteria.

Effects of O2 Concentrations and Various Haemoglobins on Respiration and Nitrogenase Activity of Bacteroids from Stem and Root Nodules of Sesbania rostrata and of the Same Bacteria from Continuous Cultures

Summary: Bacteroids from both stem and root nodules of Sesbania rostrata inoculated with Rhizobium sp. strain ORS571 were studied in the presence of the respective leghaemoglobins or soybean root nodule leghaemoglobin. Concentrations of free dissolved O2 and rates of O2 consumption and nitrogenase activity were measured during steady states in a special reaction chamber. The two types of bacteroids had high-affinity terminal oxidase systems. They were capable of respiration-coupled nitrogenase activity at concentrations below 10 nM free dissolved O2; at such concentrations the homologous leghaemoglobins were more effective than soybean leghaemoglobin because of their higher affinity for O2. S. rostrata bacteroids were less efficient in nitrogenase activity (consuming 4 to 11 mol O2 per mol of C2H2 reduced) than soybean bacteroids (02/C2H4 = 3.6) in comparable experiments, but efficiency was not affected by the source of leghaemoglobin. Similar experiments with ORS571 grown in N2-fixing continuous cultures showed that these bacteria also had high-affinity terminal oxidase systems coupled to nitrogenase, but those grown at <1 μM dissolved O2 were less efficient (O2/C2H4 >21) than those grown at 7 to 11 μM-O2 (O2/C2H4 <8). Nitrogenase activity of bacteria grown at the higher O2 concentration increased when the O2 concentration in the chamber was raised from 0.1 to 3 μM in experiments in which mammalian myoglobin replaced leghaemoglobin in the reaction solution. With bacteria grown at < 1 μM-O2, nitrogenase activity was inhibited (reversibly) by 50% after an increase from 0.1 to 1.4 μM-O2 in the reaction chamber. After changes in rates of supply of dissolved O2 there were oscillations in rates of O2 consumption before establishment of new steady states. These effects were greater in bacteria grown at 7 to 11 μM-O2 than in those grown at < 1 μM-O2, but with the latter, a sixfold increase in O2 flux produced only very small increases in concentration of dissolved O2.

Alternative cyanide- and antimycin A-insensitive respiratory system in Sporobolomyces red yeasts.

Sporobolomyces ruberrimus is insensitive to antimycin A which is a respiratory inhibitor of the cytochrome system, as cyanide is. When this red yeast was cultured in the presence of antimycin A, the growth curve showed the same pattern as that of the normal culture in the absence of it, but the growth mass was only about 70% of that of the normal culture. The antimycin A-insensitive and cyanide-insensitive respiration of Sp. ruberrimus was inhibited by pyrocatechol and salicylhydroxamic acid. Sporobolomyces red yeasts have two characteristic terminal oxidase systems; one is a cytochrome oxidase system and the other is a cyanide- and antimycin A-insensitive oxidase system. The proportions of the two respiratory systems differed among the species and strains of Sporobolomyces red yeasts examined.

The cyanide sensitivity of the residual respiration inSchizosaccharomyces japonicus

Abstract Schizosaccharomyces japonicus, a highly respiratory deficient yeast species, contains two terminal oxidase systems. One is highly sensitive to cyanide like the main terminal oxidase system of respiratory sufficient yeasts. The alternative system is hardly sensitive to cyanide like the usual terminal oxidase system of other respiratory deficient yeasts, and such as that found in respiratory sufficient yeasts besides the sensitive system. The order of magnitude of each system in Sch. japonicus is only a few μ l O 2 · (mg dry biomass) −1 · h −1 , the insensitive system having the lowest activity. As a result the alternative system may pass unnoticed. This situation may be unique among yeasts.

Energy conservation in the terminal region of the respiratory chain of the methylotrophic bacterium Methylophilus methylotrophus.

Reduced + CO minus reduced difference spectra of respiratory membranes prepared from methanol-limited cultures of Methylophilus methylotrophus confirm the presence of three CO-binding cytochromes i.e. cytochromes aa3, o and cco. The kinetics of cyanide inhibition indicate that the respiratory chain of this organism is branched at the level of cytochrome c to two major terminal oxidases, viz. cytochromes aa3 and o; cytochrome cco is probably not a physiologically significant oxidase. Determination of proton and charge translocation stoichiometries (leads to H+/O and leads to K+/O quotients) during oxidation of ascorbate-N,N,N',N'-tetramethyl-p-phenylenediamine shows that the terminal oxidase system of this organism exhibits a net inward translocation of 2e-, but no net proton translocation, when a pair of electrons are passed from cytochrome c to oxygen. The use of appropriate concentrations of cyanide to selectively inhibit cytochrome o indicates that the overall translocation stoichiometries are achieved by the two oxidases, aa3 and o, functioning similarly. These and other results suggest that methanol oxidase is organised as a simple redox arm with the methanol oxidation site and oxygen consumption site(s) on the periplasmic and cytoplasmic faces of the inner membrane respectively.

Properties of Terminal Oxidase Systems of Bacteroids from Root Nodules of Soybean and Cowpea and of N2-fixing Bacteria Grown in Continuous Culture

The respiration of bacteroids from root nodules of soybean (inoculated with Rhizobium japonicum strain CB1809) and cowpea (inoculated with Rhizobium sp. strain CB756) was studied at low concentrations of dissolved O2 in a stirred electrode chamber (range 0·1 to 10 µ m-O2) and during deoxygenation of solutions containing oxyleghaemoglobin (range 0·003 to 0·3 µ m-0·2) or oxymyoglobin (0·1 to 10 µ m-O2). The O2 affinity of terminal oxidase systems of these bacteria (measured as apparent K s) depended on the range of concentration of free, dissolved O2 in which the measurements were made. This was due to the capacity of the oxidases to express increasing affinity as the O2 concentration declined below certain transition concentrations, thus tending to maintain the respiration rate. In the lowest concentration range (0.003 to 0·01 µ m-O2) an oxidase of very high affinity for O2 (apparent K s 0·005 µ m) appeared to be under allosteric control. Three additional oxidases or oxidase affinity states could be recognized. In strain CB1809, the one with lowest affinity was insensitive to CO and less sensitive to N-phenylimidazole and azide, whilst the oxidases of higher affinity were very sensitive to these inhibitors. Terminal oxidase systems of the same strains from O2-limited continuous cultures resembled those of bacteroids, when assayed in the same way. The properties of terminal oxidases of Azotobacter vinelandii strain AVO, Azospirillum brasilense strain Sp7 and Klebsiella pneumoniae strain 50231, when grown in O2-limited continuous cultures, were examined in the same experimental systems. In all culture states examined there was no evidence for multiple oxidases as seen with the Rhizobium spp. In two of these bacteria, Azospirillum brasilense and K. pneumoniae, the oxidases appeared to be allosteric but their affinities were very different (apparent K s 0·006 and 0·11 µ m, respectively). The terminal oxidase of Azotobacter vinelandii obeyed Michaelis-Menten kinetics, but had a lower affinity for O2 (apparent K s 0·48 µ m).

An inducible n-alkane hydroxylase system containing cytochrome "O" from Candida lipolytica.

In this work the terminal oxidase system of C. lipolytica grown on n-alkanes was identified, and partially purified. Spectral characteristics typical of cytochrome O are obtained, inhibition and photodissociation of CO are reported.

Leghaemoglobin and the supply of O2 to nitrogen-fixing root nodule bacteroids: presence of two oxidase systems and ATP production at low free O2 concentration.

Studies of rates of consumption of dissolved O2 by suspensions of bacteroids (Rhizobium japonicum, strain CB1809) from soybean root nodules showed the presence of two different terminal oxidase systems. A high-affinity system, sensitive to inhibition by N-phenylimidazole and by carbon monoxide, was most active when the dissolved O2 was between 0-01 and 0-1 muM. At 1 muM-O2 or higher, this oxidase system had little activity and O2 was consumed largely by a low-affinity system insensitive to these inhibitors. At low concentrations of dissolved O2, bacteroid respiration rates appeared to be diffusion-limited. When purified oxyleghaemoglobin was added to such systems, this restriction was relieved and respiration was maintained to much lower concentrations of free dissolved O2, where nitrogenase activity was greatest. Analysis of reactions which were terminated at various stages during the depletion of O2 from oxyleghaemoglobin showed that at low free O2 concentration, the high-affinity pathway produced up to five times greater bacteroid ATP concentrations than the low-affinity oxidase pathway operating about 1 muM free O2 in the absence of leghaemoglobin. At intermediate free O2 concentrations, occurring during the later stages of deoxygenation of oxymyoglobin, intermediate concentrations of ATP were found in the bacteroids.

Muscle tricarboxylic acid cycle in Ascaris lumbricoides var. suis

Muscle homogenates of Ascaris lumbricoides var. suis metabolized succinate-2,3-C 14, acetate-1-C 14 and citrate-1,5-C 14 resulting in the formation of C 14O 2. When muscle strips were used, C 14O 2 formation from succinate-2,3-C 14 and from acetate-1-C 14 was inhibited considerably by monofluoroacetate. The incubation of succinate-2,3-C 14 or acetate-2-C 14 with particle fractions from Ascaris muscle produced C 14-citrate. Also the activities of isocitrate dehydrogenase (NADP specific), fumarase, α-Ketoglutarate dehydrogenase, malate dehydrogenase, aconitase, condensing enzyme and aceto-CoA kinase were detected in various preparations from Ascaris muscle by appropriate assay methods. These findings suggested the occurrence of a tricarboxylic acid cycle in Ascaris muscle. On the other hand, examination of C 14-labeled succinate and fumarate catabolism has revealed that the carboxylic acid metabolism in Ascaris muscle could proceed through a dismutation reaction between carboxylic acids of different oxidation levels. The functioning of the tricarboxylic acid cycle in Ascaris muscle appeared to be considerably limited on account of the relatively low activity of terminal oxidase system as well as aconitase, condensing enzyme and pyruvate dehydrogenase. Therefore the physiological role of the cycle might be of a minor importance as compared to the dismutation type reaction.

LACTIC DEHYDROGENASES OF PSEUDOMONAS NATRIEGENS.

Walker, Hazel (University of Georgia, Athens), and R. G. Eagon. Lactic dehydrogenases of Pseudomonas natriegens. J. Bacteriol. 88:25-30. 1964.-Lactic dehydrogenases specific for d- and l-lactate were demonstrated in Pseudomonas natriegens. The l-lactic dehydrogenase showed considerable heat stability, and 40% of the activity remained in extracts after heating at 60 C for 10 min. An essential thiol group for enzyme activity was noted. The results of these experiments were consistent with the view that lactate was dehydrogenated initially by a flavin cofactor and that electrons were transported through a complete terminal oxidase system to oxygen. The intracellular site of these lactic dehydrogenases was shown to be the cell membrane. It was suggested that the main physiological role of these lactic dehydrogenases is that of lactate utilization.

STUDIES ON THE RESPIRATION OF SEA-URCHIN SPERMATOZOA

1. The activity of cytochrome oxidase in sea-urchin spermatozoa ( Hemicentrotus pulcherrimus ) was studied in relation to the Dilution Effect, using dimethyl- p -phenylene- diamine (dpphd), p -phenylenediamine (pphd), hydroquinone, ascorbic acid and cysteine as substrates. 2. All substrates except hydroquinone cause a marked rise in oxygen uptake in dense sperm suspensions (1/20). It is shown that the maximum rate obtained with sufficient amount of dpphd in dense suspensions is almost comparable with the normal respiratory rate in dilute suspensions (1/200). The oxygen uptake of dilute suspensions is not affected by the addition of dpphd. 3. The increase in oxygen uptake caused by dpphd and pphd is completely eliminated by sodium azide (10-2 M). 4. From the results of 2 and 3, it can be inferred that cytochrome oxidase exhibits its maximum activity only in dilute suspension, probably as a result of substrate saturation. 5. Heavy metals, copper and zinc, do not seem to affect the terminal oxidase system directly, but some other part of the oxidation chain, such as the soluble SH groups. 6. The effect of cysteine is rather complicated. In the range from 10-4 to 10-3 M, causes a reduction in the initial respiratory rate as already observed with other amino-acids, but above this range, it exerts a marked stimulatory effect on oxygen uptake.

PYRIDINE NUCLEOTIDE-LINKED REACTIONS OF PSEUDOMONAS NATRIEGENS

Eagon, R. G. (University of Georgia, Athens). Pyridine nucleotide-linked reactions of Pseudomonas natriegens. J. Bacteriol. 84:819-821. 1962-The observation that Pseudomonas natriegens utilizes the Embden-Meyerhof pathway and the hexose monophosphate-pentose cycle only very slightly, even though the necessary enzymes are present, was explained by the existence of a sluggish system for the oxidation of reduced triphosphopyridine nucleotide (TPNH). Pyridine nucleotide transhydrogenase could not be detected in cell-free extracts. A very active system for the oxidation of reduced diphosphopyridine nucleotide (DPNH) was observed. Thus, since lactic acid is a major end product of glucose dissimilation and since the lactic dehydrogenase of P. natriegens does not utilize DPNH as cofactor, the Embden-Meyerhof pathway apparently operates aerobically by direct oxidation of DPNH, presumably by coupling with the terminal oxidase system rather than by coupling to synthetic reactions requiring DPNH as cofactor. A TPNH-specific glutathione reductase was detected which was inhibited by adenosine-2'-monophosphate.

CARROT TISSUE AND ETHANOL

SummaryThe presence of alcohol in an aerated external medium increases the respiration rate of carrot discs. If the alcohol is labelled no labelling appears in the CO2 emitted or the acids formed in the tissue. Alcohol dehydrogenase is shown to be located almost wholly among the soluble proteins of the cytoplasm. The failure to oxidize alcohol is attributed to its spatial separation from the terminal oxidase system located in the mitochondria.

On the Changes of the Terminal Oxidase Systems with Special Reference to the Dormancy of Potato Tubers

The present investigation was carried out in order to obtain further information as to whether some respiratory enzyme system in potato tubers of which activity is inhibited by the application of diethyldithiocarbamate (Dieca) and salcylaldoxime, may play any essential participation in the dormancy of potato tubers or not and also whether any difference of respiratory system may exist between the respiration of the dormant tuber and that of the sprouting tuber or not. The results obtained may be summarized as follows. The inhibitions of O2-uptake by the tubers due to the treatments with Dieca or salcylaldoxime were reduced if the tubers were stored previously at low temperature or the dormant tubers were treated previously with gibberellin or thiourea in order to hasten their sprouting. A similar situation was found to be true in the case of the secondary-growth tuber. Judging from the results stated above, it seems quite reasonable to assume that polyphenol oxidase may take part in the respiratory enzyme systems in the dormant tubers and this polyphenol oxidase may play an important role in the dormancy of the potato tubers. The inhibiting effects of Dieca and salcylaldoxime on the O2-uptake of the potato tubers were reduced if when use was made of tubers at their sprouting time and of tubers infected with witches, broom virus which usually show no dormancy. Although the respiration rates of these tubers decreased also under the condition of low oxygen partial pressure of 5 %, a slight respiratory inhibition by the treatment with Antimycin A was recognized. Accordingly, it seems highly probable that some respiratory enzyme system other than polyphenol oxidase may exist in these tubers, but no adequate explanation can be given on the nature of this enzyme system at present. In conclusion, the copper-containing enzyme system, highly presumably polyphenol oxidase and another respiratory enzyme system which is sensitive to low oxygen partial pressure and is not yet identified as to its nature, may play physiologically important roles in dormancy of the potato tubers.

海産二枚貝組織の末端酸化系について

In a previous paper (KAWAI, '56), the evidence was presented that the respiration of gill of marine lamellibranchs, such as the oyster (Crassostrea gigas) and the pearl oyster (Pinctada martensii), is mediated through the cytochrome system. Recently, JODREY and WILBUR ('55) suggested that the cytochrome system may not play a major role in the oxidation in the mantle of oyster. It therefore seemed appropriate to establish whether the cytochrome system functions or not as the terminal oxidase system in marine lamellibranch tissues other than the gill. This paper concerns the effects of inhibitors on the endogeneous respiration of various tissues of the oyster, the pearl oyster and the mussel (Mytilus crassitesta), and the identification of the cytochromes in the tissues. 1. The respiration was strongly inhibited by 10-3 M cyanide, and the inhibition was partially reversed by 6×10-5 M methylene blue (Fig. 1 & 2). The inhibition degree of pearl oyster tissues was not so large compared with that of oyster. 2. In the mixture of 90% CO and 10% 02, the oxygen uptake of lamellibranch tissues was inhibited by about 50% in the dark, and the inhibition was completely reversed by light (Fig. 3, 4 & 5). 3. The reduced band of cytochrome b was detected in all tissues dexamined wheras the band a detected only in certain tissues by the spectroscopic examination (Table 1). The band c, however, could not be detected in lamellibranch tissues. A very strong pyridine hemochromogen band (560mμ), which is considered as the band of cytochrome b pyridine hemochrome, was readily obtained in every tissue examined. 4. These results lead to the conclusion that the respiration of marine lamellibranch tissues other than the gill is also mediated through the cytochrome system.

Histochemical studies on the lesions of rice blast caused by Piricularia oryzae CAV.

This paper deals with the changes in the respiration of leaf tissues infected by Piricularia oryzae. Rice varieties, Aiti-asahi (susceptible) and Kanto 53 (resistant) were used. Leaf lesions occurring on these varieties were classified into 5 types as shown diagramatically in Fig. 1. Lesions of the types 3, 4 and 5 were used for the experiments. Types 4 and represent the lesions on the susceptible var5ety and type 3 represents the lesion on the resistant variety. From each lesion, the central necrotic area is removed and a marginal zone of about 3mm width was taken as the material for measurement of the respiration rate. If the tissue is resistant, the respiration is stimulated to some extent, not exeeding over two times as that of non-infected tissue, while f the tissue is susceptible, the respiration is stimulated up to 4 to 5 times. The RQ gives usually 0.8 in the healthy, and 0.9 in the infected tissues. The diminution of Pasteur effect (shown by QairCo2/QN2Co2) is remarkeble in the susceptible tissue, but in the resistant tissue it is not affected. The respiration of resistant tissue is inhibited by 10-25 per cent by such metal enzyme inhibitors as NaN3, 8-hydroxyquinoline, and salicylaldoxime, while, that of susceptible tissue is not affected, suggesting that flavoprotein enzyme works as terminal oxidase in the infected tissue, in place of metal protein enzymes. The above mentioned change of terminal oxidase system in the infected tissue is probable because it is already known that P. oryzae produces two toxins, α-picolinic acid. and piricularin and that these are inhibitory to metal enzymes (Tamari et al.). The writers consider that the main cause of unusual stimulation of respiration and the diminution of Pasteur effect in susceptible tissue is the change of terminal oxidase, from metal protein to flavoprotein.

STUDIES ON THE RESPIRATORY SYSTEM OF THE POKY STRAIN OF NEUROSPORA

It was shown recently that the poky strain of Neurospora accumulates large amounts of cytochrome c, but is deficient in cytochromes a and b and thus in cytochrome oxidase and succinic acid oxidase activities (1). These findings, which suggest that respiration in poky does not utilize the cytochrome system, led to a study of the respiratory system of poky compared with that of wild type. The experiments reported here are concerned with the effects of the inhibitors cyanide and azide on the respiration of intact mycelium of poky and wild type, the oxygen uptake of cell-free extracts in the oxidation of their endogenous substrates, the cofactor requirement for this oxidation, and the effects of azide and of oxygen tension. Evidence presented confirms that, in poky, respiration does not depend upon the cytochromes, but upon another terminal oxidase system in which flavin-adenine dinucleotide (FAD) enzymes appear to be concerned.