Demonstration of Physical Interactions between Consecutive Enzymes of the Citric Acid Cycle and of the Aspartate‐Malate Shuttle

Beeckmans, Sonia; Kanarek, Louis

doi:10.1111/j.1432-1033.1981.tb06369.x

Cited by 132 publications

(30 citation statements)

References 67 publications

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“…Enzymes of the TCA cycle have been shown to physically interact with MAS enzymes, allowing for direct communication of metabolic changes between the mitochondrial matrix and the cytosol (27). The TCA cycle is thought to function similarly in zebrafish and humans (28, 29).…”

Section: Discussionmentioning

confidence: 99%

Visnagin protects against doxorubicin-induced cardiomyopathy through modulation of mitochondrial malate dehydrogenase

et al. 2014

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Doxorubicin is a highly effective anti-cancer chemotherapy agent, but its usage is limited by its cardiotoxicity. To develop a drug that prevents the cardiac toxicity of doxorubicin while preserving its anti-tumor potency, we established a doxorubicin-induced cardiomyopathy model in zebrafish that recapitulated the cardiomyocyte apoptosis and contractility decline observed in patients. Using this model, we screened 3000 compounds and discovered that visnagin (VIS) and diphenylurea (DPU) rescue cardiac performance and circulatory defects caused by doxorubicin treatment in zebrafish. VIS and DPU reduced doxorubicin-induced apoptosis in cultured cardiomyocytes and in vivo in zebrafish and mouse hearts. Furthermore, VIS treatment improved cardiac contractility in doxorubicin-treated mice. Importantly, VIS and DPU caused no reduction in the chemotherapeutic efficacy of doxorubicin in several cultured tumor lines or in zebrafish and mouse xenograft models. Using affinity chromatography, we discovered that VIS binds to mitochondrial malate dehydrogenase (MDH2), one of the key enzymes in the tricarboxylic acid cycle. As with VIS, treatment with the MDH2 inhibitors mebendazole, thyroxine, and iodine prevented doxorubicin cardiotoxicity, as did treatment with malate itself, suggesting that modulation of MDH2 activity is responsible for VIS’s cardioprotective effects. Taken together, this study identified VIS and DPU as potent cardioprotective compounds and implicates MDH2 as a previously undescribed, druggable target for doxorubicin-induced cardiomyopathy.

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Section: Discussionmentioning

confidence: 99%

Visnagin protects against doxorubicin-induced cardiomyopathy through modulation of mitochondrial malate dehydrogenase

et al. 2014

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“…Similar to the tricarboxylic acid metabolon, we anticipated that purine biosynthetic enzyme activity in the purinosome could be enhanced by concentrating the proteins within a smaller volume to better sense metabolite levels and coordinate their cellular movements [56-59]. These precedents led us to investigate the locus of the purinosome within a cell.…”

Section: Purinosome Functionmentioning

confidence: 99%

A New View into the Regulation of Purine Metabolism: The Purinosome

Pedley

Benkovic

2017

Trends in Biochemical Sciences

398

361

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Other than serving as building blocks for DNA and RNA, purines metabolites provide a cell with the necessary energy and cofactors to promote cell survival and proliferation. A renewed interest in how purine metabolism may fuel cancer progression has uncovered a new perspective into how a cell regulates purine need. Under cellular conditions of high purine demand, the de novo purine biosynthetic enzymes cluster near mitochondria and microtubules to form dynamic multi-enzyme complexes referred to as purinosomes. This review highlights the purinosome as a novel level of metabolic organization of enzymes in cells, its consequences for regulation of purine metabolism, and the extent that purine metabolism is being targeted for the treatment of cancers.

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“…It is therefore likely that these chaperones also played a vital role in the refolding of the recombinant T. emersonii m-MDH to yield active enzyme. As m-MDH is a constitutively expressed enzyme in both prokaryotic and eukaryotic systems, Western blot analysis using an anti-His 6 Ig was used to confirm that the purified recombinant protein (Fig. 4B), was the His-tagged m-MDH from T. emersonii revealing a single band of the correct molecular mass (35 kDa).…”

Section: Expression Of Recombinant Malate Dehydrogenase In E Colimentioning

confidence: 99%

“…Investigation of the optimum pH for m-MDH activity in the oxaloacetate reductase direction was performed with 200 lM oxaloacetate and 200 lM NADH over a wide pH range (4)(5)(6)(7)(8)(9)(10)(11)(12) and at 30°C. Determination of the pH optimum in the malate oxidation direction was achieved with 10 mM malate and 1 mM NAD + as substrate and cosubstrate, respectively, over the same pH range at 30°C.…”

Section: Determination Of Selected Biochemical Propertiesmentioning

confidence: 99%

“…Mitochondrial MDH displays a complex regulatory pattern that involves allosteric activation [2], membrane interaction [3] and formation of multienzyme complexes [4] with enzymes such as citrate synthase, aspartate aminotransferase and other mitochondrial components [5][6][7][8][9][10]. Much of the previous research has focused on the comparison of primary and three-dimensional structures of mitochondrial and cytoplasmic forms of MDH isolated from a single source, e.g.…”

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confidence: 99%

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Mitochondrial malate dehydrogenase from the thermophilic, filamentous fungus Talaromyces emersonii

Maloney

Callan

Murray

et al. 2004

European Journal of Biochemistry

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Mitochondrial malate dehydrogenase (m-MDH; EC 1.1.1.37), from mycelial extracts of the thermophilic, aerobic fungus Talaromyces emersonii, was purified to homogeneity by sequential hydrophobic interaction and biospecific affinity chromatography steps. Native m-MDH was a dimer with an apparent monomer mass of 35 kDa and was most active at pH 7.5 and 52°C in the oxaloacetate reductase direction. Substrate specificity and kinetic studies demonstrated the strict specificity of this enzyme, and its closer similarity to vertebrate m-MDHs than homologs from invertebrate or mesophilic fungal sources. The fulllength m-MDH gene and its corresponding cDNA were cloned using degenerate primers derived from the N-terminal amino acid sequence of the native protein and multiple sequence alignments from conserved regions of other m-MDH genes. The m-MDH gene is the first oxidoreductase gene cloned from T. emersonii and is the first full-length m-MDH gene isolated from a filamentous fungal species and a thermophilic eukaryote. Recombinant m-MDH was expressed in Escherichia coli, as a His-tagged protein and was purified to apparent homogeneity by metal chelate chromatography on an Ni 2+ -nitrilotriacetic acid matrix, at a yield of 250 mg pure protein per liter of culture. The recombinant enzyme behaved as a dimer under nondenaturing conditions. Expression of the recombinant protein was confirmed by Western blot analysis using an antibody against the His-tag. Thermal stability studies were performed with the recombinant protein to investigate if results were consistent with those obtained for the native enzyme.Keywords: malate dehydrogenase; filamentous fungus; bio-specific affinity chromatography; thermophilic eukaryote; Talaromyces emersonii.Malate dehydrogenase (MDH) catalyzes the pyridine nucleotide-dependent interconversion of malate and oxaloacetate in the tricarboxylic acid cycle. It is also thought to have a role in the malate/aspartate shuttle across the inner mitochondrial membrane. In most eukaryotic cells there are two major isoenzymes of MDH, cytosolic MDH (c-MDH) and mitochondrial MDH (m-MDH) [1].The majority of m-MDHs isolated to date from eukaryotic sources are homodimers of identical subunits, with a monomer size of 34 kDa. Mitochondrial MDH displays a complex regulatory pattern that involves allosteric activation [2], membrane interaction [3] and formation of multienzyme complexes [4] with enzymes such as citrate synthase, aspartate aminotransferase and other mitochondrial components [5][6][7][8][9][10]. Much of the previous research has focused on the comparison of primary and three-dimensional structures of mitochondrial and cytoplasmic forms of MDH isolated from a single source, e.g. pig heart tissue [11,12]. In contrast to bacterial, vertebrate and invertebrate m-MDH, a paucity of information exists on fungal m-MDH with significantly more known about yeast m-MDH than homologs from filamentous fungal species. Previous research by Dalhaus et al. [13] and Goward and Nicholls [14] has proposed that investigation...

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Demonstration of Physical Interactions between Consecutive Enzymes of the Citric Acid Cycle and of the Aspartate‐Malate Shuttle

Cited by 132 publications

References 67 publications

Visnagin protects against doxorubicin-induced cardiomyopathy through modulation of mitochondrial malate dehydrogenase

Visnagin protects against doxorubicin-induced cardiomyopathy through modulation of mitochondrial malate dehydrogenase

A New View into the Regulation of Purine Metabolism: The Purinosome

Mitochondrial malate dehydrogenase from the thermophilic, filamentous fungus Talaromyces emersonii

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