The presence of a short redox chain in the membrane of intact potato tuber peroxisomes and the association of malate dehydrogenase with the peroxisomal membrane

Struglics, A.; Fredlund, Kenneth M.; Rasmusson, Allan G.

doi:10.1034/j.1399-3054.1993.880103.x

Cited by 22 publications

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“…Although POM only contain a maximum of 0.2% peroxisomes on a protein basis, the catalases could be contaminants, since catalase makes up approximately 50% of all the peroxisomal protein in potato tubers (Struglics et al, 1993). However, catalase has been reported to be present inside rat heart mitochondria (Radi et al, 1991), so the parsimonious explanation of catalase inside plant mitochondria cannot be excluded.…”

Section: Turnover Of Rosmentioning

confidence: 90%

“…It has previously been established that very pure mitochondria can be obtained from potato tubers, which only contain 0.2% peroxisomes or plastids on a protein basis (Neuburger et al, 1982;Struglics et al, 1993;Considine et al, 2003). We employed this established procedure for the isolation of mitochondria with a recognized low contamination level.…”

Section: The Isolated Pom Are Highly Purifiedmentioning

confidence: 99%

“…The potato tuber is a large, relatively homogenous storage organ, and POM are therefore expected to be similarly homogenous. There are well-established methods for isolating these mitochondria in a purified, intact, and functional form (Neuburger et al, 1982;Struglics et al, 1993;Considine et al, 2003), and their basic properties are therefore well characterized. In addition, the potato genome has recently been sequenced (Xu et al, 2011), which greatly simplifies the task of identifying potato proteins.…”

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The Potato Tuber Mitochondrial Proteome

et al. 2013

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Mitochondria are called the powerhouses of the cell. To better understand the role of mitochondria in maintaining and regulating metabolism in storage tissues, highly purified mitochondria were isolated from dormant potato tubers (Solanum tuberosum 'Folva') and their proteome investigated. Proteins were resolved by one-dimensional gel electrophoresis, and tryptic peptides were extracted from gel slices and analyzed by liquid chromatography-tandem mass spectrometry using an Orbitrap XL. Using four different search programs, a total of 1,060 nonredundant proteins were identified in a quantitative manner using normalized spectral counts including as many as 5-fold more "extreme" proteins (low mass, high isoelectric point, hydrophobic) than previous mitochondrial proteome studies. We estimate that this compendium of proteins represents a high coverage of the potato tuber mitochondrial proteome (possibly as high as 85%). The dynamic range of protein expression spanned 1,800-fold and included nearly all components of the electron transport chain, tricarboxylic acid cycle, and protein import apparatus. Additionally, we identified 71 pentatricopeptide repeat proteins, 29 membrane carriers/transporters, a number of new proteins involved in coenzyme biosynthesis and iron metabolism, the pyruvate dehydrogenase kinase, and a type 2C protein phosphatase that may catalyze the dephosphorylation of the pyruvate dehydrogenase complex. Systematic analysis of prominent posttranslational modifications revealed that more than 50% of the identified proteins harbor at least one modification. The most prominently observed class of posttranslational modifications was oxidative modifications. This study reveals approximately 500 new or previously unconfirmed plant mitochondrial proteins and outlines a facile strategy for unbiased, near-comprehensive identification of mitochondrial proteins and their modified forms.

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Section: Turnover Of Rosmentioning

confidence: 90%

Section: The Isolated Pom Are Highly Purifiedmentioning

confidence: 99%

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The Potato Tuber Mitochondrial Proteome

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“…Mitochondria were prepared from potato (Solanum tuberosum L.) tubers according to [16]. Crude mitochondria were isolated from Arum maculatum spadices according to [17] and purified according to [18].…”

Section: Methodsmentioning

confidence: 99%

Direct evidence for the presence of two external NAD(P)H dehydrogenases coupled to the electron transport chain in plant mitochondria

1995

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Exogenous NADPH oxidation by purified mitochondria from both potato tuber and Arum maculatum spadix was completely and irreversibly inhibited by sub-micromolar diphen~leneiodonium (DPI), while exogenous NADH oxidation was inhibited to only a small degree. Addition of DPI caused the collapse of the membrane potential generated by NADPH oxidation, while the potential generated by NADH was unaffected. We conclude that there are two distinct enzymes on the outer surface of the inner membrane of plant mitochondria, one specific for NADH, the other relatively specific for NADPH, with both enzymes linked to the electron transport chain.Key words: Diphenyleneiodonium; Electron transport chain; Mitochondria, plant; NAD(P)H dehydrogenase |. IntroductionUnlike mammalian mitochondria, the electron transport hain of plant mitochondria contains at least two NAD(P)H ,tehydrogenases in addition to Complex I [1]. One of these ~otenone-insensitive dehydrogenases is located on the outer , urface of the inner mitochondrial membrane, the other on the inner, matrix surface, and in the following we will refer to them ;s NDex and NDin, respectively.Due to the presence of NDex, plant mitochondria oxidize ~xogenous NADH and NADPH. The resulting electrons enter he electron transport chain at ubiquinone bypassing Complex i, the site of rotenone inhibition [1][2][3][4][5]. Compared to NADH ~xidation, NADPH oxidation has a lower pH optimum [6][7][8], s more sensitive to thiol reagents [7,9] and appears to require nore Ca 2÷ for activity [9][10][11][12]. NADH oxidation has been re~orted to be induced in red beetroots without a concomitant nduction of NADPH oxidation [13]. Likewise, mitochondria rom suspension-cultured cells of sugar beet oxidize NADH, ~ut exhibit very low rates of NADPH oxidation [14]. On the )asis of this circumstantial evidence it has been concluded that t is most likely that two separate enzymes exist, one for each ,:oenzyme [2,4,14]. However, no direct evidence has ever been r~resented to support this conclusion.Diphenyleneiodonium ( Materials and methodsMitochondria were prepared from potato (Solanum tuberosum L.) tubers according to [16]. Crude mitochondria were isolated from Arum maculatum spadices according to [17] and purified according to [18].Oxygen consumption was measured in a medium containing 0.3 M sucrose, 5 mM MOPS (pH 7.2), 5 mM KH:PO4, 2.5 mM MgCI2, 1 mM CaC12 and 0.4 ¢tM FCCP using an oxygen electrode (Rank Bros.). The membrane of the oxygen electrode was washed with 70% (v/v) ethanol between assays to avoid carry-over of DPI.Membrane potential measurements using 16 ]zM safranine as the probe were made according to [19]. The medium used was the same as that above except that FCCP was omitted, CaCI2 was 0.1 mM and 20 ,ug.mg 1 BSA was included.NAD(P)H oxidation with oxygen as the electron acceptor was assayed spectrophotometrically at 340 nm in the same medium used for the oxygen electrode measurements. Activities were calculated using an extinction coefficient for NAD(P)H of 6.2 mM -1 .cm -~, All ...

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“…Mitochondria and peroxisomes were isolated from pea leaves by differential and density-gradient centrifugation, as described by Struglics et al (1993), but with some minor modifications. Leaves (40 g) were chopped with a polytron (Kinematica Krienz, Luzern, Switzerland) into a medium containing 30 mM Mops (pH 7.3), 3 mM EDTA, 25 mM Cys, 0.3 M mannitol, and 0.3% ( w / v ) BSA, using 4 volumes of medium 8-l fresh weight, and the homogenates were filtered through two layers of Miracloth (Calbiochem).…”

Section: Purification Of Cell Organellesmentioning

confidence: 99%

Evidence for the Presence of the Ascorbate-Glutathione Cycle in Mitochondria and Peroxisomes of Pea Leaves

et al. 1997

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l h e presence of the enzymes of the ascorbate-glutathione cycle was investigated in mitochondria and peroxisomes purified from pea (Pisum sativum L.) leaves. AI1 four enzymes, ascorbate peroxidase (APX; EC 1.1 1.1.1 l ) , monodehydroascorbate reductase (EC 1.6.5.4), dehydroascorbate reductase (EC 1.8.5.1), and glutathione reductase (EC 1.6.4.2), were present in mitochondria and peroxisomes, as well as i n the antioxidants ascorbate and glutathione. l h e activity of the ascorbate-glutathione cycle enzymes was higher in mitochondria than in peroxisomes, except for APX, which was more active in peroxisomes than in mitochondria. lntact mitochondria and peroxisomes had no latent APX activity, and this remained i n the membrane fraction after solubilization assays with 0.2 M KCI.Monodehydroascorbate reductase was highly latent in intact mitochondria and peroxisomes and was membrane-bound, suggesting that the electron acceptor and donor sites of this redox protein are not on the externa1 side of the mitochondrial and peroxisomal membranes. Dehydroascorbate reductase was found mainly in the soluble peroxisomal and mitochondrial fractions. Clutathione reductase had a high latency in mitochondria and peroxisomes and was present i n the soluble fractions of both organelles. I n intact peroxisomes and mitochondria, the presence of reduced ascorbate and glutathione and the oxidized forms of ascorbate and glutathione were demonstrated by high-performance liquid chromatography analysis. l h e ascorbate-glutathione cycle of mitochondria and peroxisomes could represent an important antioxidant protection system against H,O, generated in both plant organelles.

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The presence of a short redox chain in the membrane of intact potato tuber peroxisomes and the association of malate dehydrogenase with the peroxisomal membrane

Cited by 22 publications

References 0 publications

The Potato Tuber Mitochondrial Proteome

The Potato Tuber Mitochondrial Proteome

Direct evidence for the presence of two external NAD(P)H dehydrogenases coupled to the electron transport chain in plant mitochondria

Evidence for the Presence of the Ascorbate-Glutathione Cycle in Mitochondria and Peroxisomes of Pea Leaves

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