Properties of Higher Plant Mitochondria. III. Effects of Respiratory Inhibitors

Ikuma, Hiroshi; Bonner, Walter D.

doi:10.1104/pp.42.11.1535

Cited by 78 publications

(72 citation statements)

References 28 publications

(36 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Rotenone and amytal have been used as inhibitors of electron transport in plant mitochondria, however, only incomplete inhibition has been observed [21]. I n this study piericidin A, an inhibitor of electron transport in the region of the iron-sulphur proteins of the NADH dehydrogenase [22] has been employed to investigate the NADH dehydrogenase responsible for the oxidation of pyridine nucleotide reduced by different substrates.…”

Section: Piericidin a Sensitivity And Location Of The Pyridine-nucleomentioning

confidence: 99%

Pathways for the Oxidation of Malate and Reduced Pyridine Nucleotide by Wheat Mitochondria

Brunton

Palmer

1973

European Journal of Biochemistry

View full text Add to dashboard Cite

piericidin A and oxaloacetate have been used as inhibitors to investigate the operation and localisation of NAD+-linked dehydrogenases and the possible compartmentation of these enzymes within the matrix of the mitochondrion. 1.Mitochondria isolated from etiolated shoots of wheat (Triticum vulgare) were shown to oxidise a variety of NAD+-linked substrates. Oxaloacetate, when added to rapidly respiring preparations (i.e. state 3) resulted in a large but transient inhibition ofthe oxidation of both pyruvate and citrate, but had little effect on the rate of oxidation of malate.2. The products of oxidation of malate, in the absence of added NAD+, were found to be a large quantity of pyruvate together with smaller quantities of oxaloacetate and citrate. Using potassium ferricyanide as a non-penetrating electron acceptor it was possible to show that the malate oxidation occurred in the matrix. This is consistent with the operation of both the malic enzyme and a low level of malate dehydrogenase within the matrix of the mitochondrion.3. The low level of inhibition of malate oxidation by oxaloacetate was a function of the concentration of malate and appeared to be of a competitive nature.4. Measurement of the redox state of the endogenous NAD+ pool showed that under phosphorylatig conditions both malate and citrate reduce approximately 15O/, of the total coenzyme. The addition of 0.2 mM oxaloacetate to phosphorylating aerobic mitochondria failed to oxidise any of the NAD+ reduced by malate but resulted in a partial oxidation of NAD+ reduced by citrate. The transition from the aerobic to anaerobic state in the presence of malate resulted in a biphasic reduction of the endogenous NAD+, whereas in the presence of citrate it was monophasic.5 . The oxidation of NAD+-linked substrates, including malate, was found to be only partially inhibited by piercidin A. The piericidin-A-resistant pathway was found to be located internally on the basis of experiments using ferricyanide as a non-penetrating electron acceptor. The first site of phosphorylation was not coupled to oxidation mediated by the piericidin-A-resistant pathway.6. The presence of piericidin A altered the shape of the malate saturation curve. I n the ahsence of the inhibitor it was found to be biphasic; saturating at 60 mM and in its presence to be monophasic saturing a t 15 mM. At high concentrations of malate (60 mM) piericidin A caused approximately a 30 O/, inhibition, similar to that found with other NAD+-linked substrates, while a t lower concentrations of malate (15 mM) piericidin A caused a much higher inhibition (80 O/,), which subsequently recovered to the much higher piericidin-A-resistant rate observed with higher concentrations of malate. 7.I n the presence of piericidin A oxaloacetate caused severe but transient inhibition of malate oxidation, similar in magnitude to that observed during pyruvate oxidation in the absence of piericidin A.8. It was concluded that the data were consistent with the spatial separation of both the internal malic enzyme and ma...

show abstract

Section: Piericidin a Sensitivity And Location Of The Pyridine-nucleomentioning

confidence: 99%

Pathways for the Oxidation of Malate and Reduced Pyridine Nucleotide by Wheat Mitochondria

Brunton

Palmer

1973

European Journal of Biochemistry

View full text Add to dashboard Cite

show abstract

“…II the work reported here, mitochon,dria from the hvpocotyls of etiolated muing beans (Phaseolifs aurcufs) Were iused. since the method of preparation and the responses of these mitochondria to stubstrates, inhlibitors, and tincouplers have been studied inI depth bh-Ikumna and Bonner (22,23,24 (27).…”

mentioning

confidence: 99%

The Respiratory Chain of Plant Mitochondria. III. Oxidation Rates of the Cytochromes c and b in Mung Bean Mitochondria Reduced With Succinate

Storey¹

1969

Plant Physiol.

View full text Add to dashboard Cite

Abstract. Oxidation rates of the 2 cytochromes c and the 3 cytochromes b in mitochondria from the hypocotyls of etiolated mung beans (Phaseolus aureus) have been measured with a rapid mixing flow apparatus ooupled to a dual wavelength spectrophotometer. The mitochondria were depleted with uncoupler and ADP prior to starting the experiments to minimize the complications caused by energy-linked processes on electron transport. The half-time for oxidation and absorbance change at a given wavelength for the rapidly oxidized cytochromes was plotted as a function of wavelength, with 570 m,t as reference, to yield a spectrum in which absorbance maxima and half-times were correlated. The 2 cytochrome c components have oxidation half-times of 2 milliseconds for c549 and 3 milliseconds for c ,4,. ( Mitochondria from the spadix of Artumt macildaturn have a cvtochromiie 1b with a low temperature absorption maximtlm at 5.58 inL whose oxidation is not inhibited by antimvcin A (1). This component was term-eied cytochrome b, 1y Bendall and Hill (2). Bonnler (16) has proposed that b7, and b5,, are the same cvto4chrotne. Tf this is the case, the interesting lprohlelti arises as to why its oxidation is not inlhibitedi bv an'titlivcitt A\ itl -Irini 1mitochlondria, bitt is itihibite(l ill all other plant mitochotidria so far descilbed.The existetice of 2 cvtochrome c componetnts cannot be demonstrated directly in difference spectra of plant mitochondria even at 770K. Bonner (5S) solubilized 1 cytochrome c conmponent by means of 0.25 % (w/v) NaCI solution. Its redtuced-minusoxidized difference spectrum at 770K showed an absorption maximum at 547 m,u with a shoulder at 545 mx, while the component not solubilized by this process showed a maximum at 549 miu. Subse-quentlv, the solubilization process was improved by 413www.plantphysiol.org on May 9, 2018 -Published by Downloaded from

show abstract

“…The mitochondria have been characterized (6)(7)(8) with respect to a series of substrates, inhibitors, and uncouplers of electron transport and oxidative phosphorylation.…”

mentioning

confidence: 99%

Effects of Guanidine Inhibitors on Mung Bean Mitochondria

Wilson

Bonner

1970

Plant Physiol.

View full text Add to dashboard Cite

The effects of phenylethylbiguanidide, decamethylenediguanidide, and Ikuma and Bonner (6) have described a reliable method for the production of tightly coupled mitochondria from etiolated mung bean hypocotyl tissue. The mitochondria have been characterized (6-8) with respect to a series of substrates, inhibitors, and uncouplers of electron transport and oxidative phosphorylation.Extensive studies of the guanidine inhibitors have been carried out with rat liver mitochondria, in which a differential sensitivity of the three coupling sites to the various guanidine inhibitors has been observed. Coupling site 1 is most effectively inhibited by the alkylguanidines, the effectiveness of the compounds increasing with increasing chain length. Octylguanidine was found to be the most effective compound acting on site 1 (10). Chappell observed that the alkylguanidines were most effective when supplied to the rat liver mitochondria in states 1 or 4 (3). DBI2 appears to be specific in its inhibition of the coupling site 2 (11) while synthalin has been reported to be a specific inhibitor of coupling site 3 (4).1 This work was supported by a grant from the National Science Foundation.2 Abbreviations: DBI: phenylethylbiguanidide; synthalin: decamethylenediguanidide; 1799: bis(hexafluoroacetonitryl) acetone; TMPD: N,N,N1,Nl-tetramethyl-p-phenylenediamine.The guanidine inhibitors appear to have a site of action closer to the respiratory chain than either oligomycin or uncouplers of oxidative phosphorylation. Pressman has observed a slow release of the inhibition of the alkyguanidine to uncouplers sucb as DNP (11).The guanidine inhibitors have not been studied, however, with plant mitochondria. In this paper the effects of octylguanidine, DBI, and synthalin have been examined on mung bean mitochondria. Differential effects of octylguanidine on the three coupling sites and the cyanide-insensitive respiration will be described. METHODSMitochondria were prepared from the hypocotyls of 5-day-old dark-grown mung beans (Phaseolus aureus var. Jumbo) as described earlier (6). The mung bean tissue was disrupted by a 5-sec treatment with a Polytron mixer.Octylguanidine sulphate, DBI, and synthalin were kindly supplied by Dr The effects of octylguanidine on site 2 by method A was studied by following the reduction of 3 mm potassium ferricyanide (9) in the presence of 0.06 mm KCN at 420 to 470 nm with a double beam spectrophotometer or at 420 rim in a split beam spectrophotometer (2). 21www.plantphysiol.org on May 10, 2018 -Published by Downloaded from

show abstract

Properties of Higher Plant Mitochondria. III. Effects of Respiratory Inhibitors

Cited by 78 publications

References 28 publications

Pathways for the Oxidation of Malate and Reduced Pyridine Nucleotide by Wheat Mitochondria

Pathways for the Oxidation of Malate and Reduced Pyridine Nucleotide by Wheat Mitochondria

The Respiratory Chain of Plant Mitochondria. III. Oxidation Rates of the Cytochromes c and b in Mung Bean Mitochondria Reduced With Succinate

Effects of Guanidine Inhibitors on Mung Bean Mitochondria

Contact Info

Product

Resources

About