Purification and characterization of branched chain α-keto acid dehydrogenase complex of bovine kidney

Pettit, Flora H.; Yeaman, Stephen J.; Reed, Lester J.

doi:10.1073/pnas.75.10.4881

Cited by 257 publications

(171 citation statements)

References 16 publications

(11 reference statements)

Supporting

Mentioning

159

Contrasting

Order By: Relevance

“…Apicomplexans have repurposed another member of this family -the branched-chain ketoacid dehydrogenase (BCKDH) complex, usually responsible for branched-chain amino acid (BCAA) degradationfor mitochondrial conversion of pyruvate to acetyl-CoA [46]. BCKDH had been hypothesized to have PDH activity in Apicomplexa because (i) BCKDH has some PDH activity in other organisms [48][49][50],[ 1 6 _ T D $ D I F F ] and (ii) other enzymes of the BCAA degradation pathway and associated detoxification steps by the methyl-citrate cycle are lacking in Plasmodium spp., resulting in the apparent 'orphan' status of this enzyme [4,6,9,47]. Genetic ablation of the functional subunit of BCKDH (E1/) in P. berghei leads to abrogation of gametocyte maturation and ookinete production, and an almost complete halt in oocyst development, consistent with increased TCA function during this differentiation ( Figure 1) [46].…”

Section: Apicomplexan Mitochondrial Enzymes: Old Dogs New Tricksmentioning

confidence: 99%

Apicomplexan Energy Metabolism: Carbon Source Promiscuity and the Quiescence Hyperbole

Jacot

Waller

Soldati‐Favre

et al. 2016

Trends in Parasitology

View full text Add to dashboard Cite

Pages 15apicomplexan mitochondrion in energy generation has remained unclear, given the glucosereplete intracellular nature of the environmental niches of these parasites and the apparent reduction of mitochondrial metabolic pathways [4][5][6][7]. For example, apicomplexan mitochondria lack 'type I' NADH dehydrogenase (NDH, complex I of the ETC) and many apparently lack the enzymes and transporters required for fatty acid b-oxidation [8,9]. Furthermore, the pyruvate dehydrogenase complex (PDH) responsible for conversion of pyruvate into acetyl-CoA -an obligate fuel for the TCA cycle -is only present in the apicoplast [10]. Consequently, there was no obvious entry point to allow catalysis of glycolytic pyruvate by the TCA cycle [10][11][12][13][14]. Despite this, there is overwhelming evidence that apicomplexans rely on a functional and canonical TCA cycle (as reviewed [4,5,15]), and apicomplexan genome annotations indicate the presence of all enzymes of the TCA cycle. Only one clade of apicomplexans, Cryptosporidium spp., show evidence of outright loss of the TCA cycle, but these taxa retain only a highly reduced mitochondrion, the mitosome, and have also lost their apicoplast [5,8,16].This review highlights how metabolomics technologies coupled to genetic manipulation have helped in unraveling apicomplexan parasite plasticity in carbon source utilization through different life stages, revealing a complex and sometimes counter-intuitive pattern of metabolic gains, losses, and reassignments. These changes have presumably been tailored to allow these parasites to thrive within their various host niches, but may constitute some much-needed Achilles' heels in the fight against these devastating diseases. Plasmodium falciparum and the Glycolytic DeceitGlycolysis has long appeared to be the main source of ATP and NAD(P)H in the erythrocytic stages of the malaria parasites, [5,[17][18][19][20][21]. Indeed, glucose uptake in P. falciparum-infected red blood cells (RBCs) has been observed to increase 75-100-fold compared to uninfected RBC [22][23][24][25]. Up to 93% of glucose is converted directly to lactate in asexual stages [26][ 4 _ T D $ D I F F ] and is ultimately excreted into the surrounding host cell. Perturbation of glucose uptake is detrimental to parasite growth [27,28], suggesting that glycolysis is an important part of the parasite's strategy for rapid proliferation. In a manner analogous to the Warburg effect observed in highly-proliferative cancer lines and other rapidly-growing cells (e.g., yeast and bloodstream Trypanosoma spp.), Plasmodium (in erythrocytic stages) has opted for fast generation of ATP through substrate-level phosphorylation and secretion of lactate as an end-product (aerobic fermentative glycolysis), as opposed to the 'slow but efficient' mitochondrial oxidative phosphorylation and complete oxidation [29]. Reasons for this dependence on glycolytic fermentation remain unclear -after all, blood stages of Plasmodium certainly have access to oxygen -but it may be a way of avoiding excessiv...

show abstract

Section: Apicomplexan Mitochondrial Enzymes: Old Dogs New Tricksmentioning

confidence: 99%

Apicomplexan Energy Metabolism: Carbon Source Promiscuity and the Quiescence Hyperbole

Jacot

Waller

Soldati‐Favre

et al. 2016

Trends in Parasitology

View full text Add to dashboard Cite

show abstract

“…It is clear that, all other things being equal, hE1b will easily dominate the competition for binding sites on the hSBDb of BCKDC. This situation arises from the much higher affinity of hE1b for hSBDb (K d ϭ 205 nM) 5 compared with that of hE3. However, BCKDC requires bound hE3 for the efficient activity of the enzyme (5).…”

Section: Solution Nmr Studies Of the Hsbdb-he3mentioning

confidence: 99%

“…By contrast, E3 is only loosely associated with the SBDb of the mammalian BCKDC, as indicated by the absence of E3 in highly purified BCKDC preparations (5-7). The low-affinity binding of E3 to the E2b core explains the loss of most BCKDC activity during the purification of bovine BCKDC at high salt concentrations (8); BCKDC activity can be recovered by the addition of exogenous E3 to the assay mixture (5). The physiological significance of the unique weak binding of E3 to the E2b core of BCKDC is unclear.…”

mentioning

confidence: 99%

Structural and Thermodynamic Basis for Weak Interactions between Dihydrolipoamide Dehydrogenase and Subunit-binding Domain of the Branched-chain α-Ketoacid Dehydrogenase Complex

Brautigam

Wynn

Chuang

et al. 2011

Journal of Biological Chemistry

View full text Add to dashboard Cite

The purified mammalian branched-chain ␣-ketoacid dehydrogenase complex (BCKDC), which catalyzes the oxidative decarboxylation of branched-chain ␣-keto acids, is essentially devoid of the constituent dihydrolipoamide dehydrogenase component (E3). The absence of E3 is associated with the low affinity of the subunit-binding domain of human BCKDC (hSBDb) for hE3. In this work, sequence alignments of hSBDb with the E3-binding domain (E3BD) of the mammalian pyruvate dehydrogenase complex show that hSBDb has an arginine at position 118, where E3BD features an asparagine. Substitution of Arg-118 with an asparagine increases the binding affinity of the R118N hSBDb variant (designated hSBDb*) for hE3 by nearly 2 orders of magnitude. The enthalpy of the binding reaction changes from endothermic with the wild-type hSBDb to exothermic with the hSBDb* variant. This higher affinity interaction allowed the determination of the crystal structure of the hE3/hSBDb* complex to 2.4-Å resolution. The structure showed that the presence of Arg-118 poses a unique, possibly steric and/or electrostatic incompatibility that could impede E3 interactions with the wild-type hSBDb. Compared with the E3/E3BD structure, the hE3/hSBDb* structure has a smaller interfacial area. Solution NMR data corroborated the interactions of hE3 with Arg-118 and Asn-118 in wild-type hSBDb and mutant hSBDb*, respectively. The NMR results also showed that the interface between hSBDb and hE3 does not change significantly from hSBDb to hSBDb*. Taken together, our results represent a starting point for explaining the long standing enigma that the E2b core of the BCKDC binds E3 far more weakly relative to other ␣-ketoacid dehydrogenase complexes.The mitochondrial ␣-ketoacid dehydrogenase complexes include the pyruvate dehydrogenase complex (PDC), 4 the ␣-ketoglutarate dehydrogenase complex and the branched-chain ␣-ketoacid dehydrogenase complex (BCKDC) (1, 2). These macromolecular protein complexes catalyze the oxidative decarboxylation of ␣-keto acids according to Reaction 1.Each ␣-ketoacid dehydrogenase complex consists of three catalytic components: a thiamine diphosphate-dependent decarboxylase/dehydrogenase (E1), a lipoyl transacylase (E2), and dihydrolipoamide dehydrogenase (E3). The overall reaction is the sum of individual reactions catalyzed by the three enzyme components, which are linked by substrate channeling. The ␣-ketoacid dehydrogenase complexes are organized around a symmetrical oligomeric E2 core, to which the E1 and E3 components are attached via noncovalent interactions. The E1 and E2 components are specific for each ␣-ketoacid dehydrogenase complex, whereas the E3 component is common among the three multienzyme complexes. The E1p, E1k, and E1b components (with the lowercase letter corresponding to each complex), similarly the E2p, E2k, and E2b components, belong to PDC, ␣-ketoglutarate dehydrogenase complex, and BCKDC, respectively. Both E1 and E3 components bind in a mutually exclusive manner to the subunit-binding domains (SBD) from the 24-meri...

show abstract

“…the oxidative decarboxylation of branched-chain 2-oxoacids derived from branched-chain amino acids. The BCOD complex comprises multiple copies of three catalytic components : a thiamine pyrophosphate-dependent branched-chain 2-oxoacid decarboxylase (E1) that contains two E1α and two E1β subunits ; a dihydrolipoyl transacylase (E2) ; and a dihydrolipoyl dehydrogenase (E3) [1,2]. In addition, the complex has a set of regulatory enzymes : a specific kinase [3] and a specific phosphatase [4], which control enzyme activity through reversible phosphorylation (inactivation)\dephosphorylation (activation) of E1 [5].…”

Section: Introductionmentioning

confidence: 99%

“…Injection of 6α-methylprednisolone (a synthetic glucoAbbreviations used : BCOD, branched-chain 2-oxoacid dehydrogenase ; DMEM, Dulbecco's modified Eagle's medium ; E1, branched-chain 2-oxoacid decarboxylase ; GAPDH, glyceraldehyde-3-phosphate dehydrogenase ; GRE, glucocorticoid-responsive element ; PEPCK, phosphoenolpyruvate carboxykinase ; PPAR, peroxisome-proliferator-activated receptor ; TAT, tyrosine aminotransferase. 1 To whom correspondence should be addressed (e-mail Chuang01!utsw.swmed.edu).…”

Section: Introductionmentioning

confidence: 99%

Down-regulation of rat mitochondrial branched-chain 2-oxoacid dehydrogenase kinase gene expression by glucocorticoids

Huang

Chuang²

1999

Biochemical Journal

View full text Add to dashboard Cite

The mammalian mitochondrial branched-chain 2-oxoacid dehydrogenase (BCOD) complex is regulated by a reversible phosphorylation (inactivation)\dephosphorylation (activation) cycle. In the present study, the effects of glucocorticoids on the level of BCOD kinase mRNA were investigated in rat hepatoma cell lines (H4IIE and FTO-2B), as well as in the rat. In H4IIE cells, dexamethasone was found to significantly reduce steadystate concentrations of BCOD kinase mRNA after a 48 h culture, and this was correlated with a 2-fold increase in the dephosphorylated form of the BCOD complex. The half-life of the kinase mRNA in H4IIE cells was not affected by dexamethasone treatment. Therefore, the decrease in the steady-state kinase mRNA level resulting from dexamethasone treatment was not caused by changes in mRNA stability, which raised the possibility of regulation at the level of gene transcription. To identify the negative glucocorticoid-responsive element in the kinase pro-

show abstract

Purification and characterization of branched chain α-keto acid dehydrogenase complex of bovine kidney

Cited by 257 publications

References 16 publications

Apicomplexan Energy Metabolism: Carbon Source Promiscuity and the Quiescence Hyperbole

Apicomplexan Energy Metabolism: Carbon Source Promiscuity and the Quiescence Hyperbole

Structural and Thermodynamic Basis for Weak Interactions between Dihydrolipoamide Dehydrogenase and Subunit-binding Domain of the Branched-chain α-Ketoacid Dehydrogenase Complex

Down-regulation of rat mitochondrial branched-chain 2-oxoacid dehydrogenase kinase gene expression by glucocorticoids

Contact Info

Product

Resources

About