Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.8.1.4 (diaphorase)
2,754 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The activity of alpha-ketoglutarate dehydrogenase complex from pigeon breast muscle is controlled by ADP and the reaction products, i. e. succinyl-CoA and NADH. ADP activates the alpha-ketoglutarate dehydrogenase component of the complex, whereas NADH inhibits alpha-ketoglutarate dehydrogenase and lipoyl dehydrogenase. In the presence of NADH the kinetic curve of the complex with respect to alpha-ketoglutarate and NAD and the dependence of upsilon versus [NAD] and upsilon versus [Lip (SH)2] in the lipoyl dehydrogenase reaction are S-shaped. In the absence of inhibitor ADP had no activating effect on lipoyl dehydrogenase; however, in the presence of NADH ADP decreases the cooperativity for NAD. The cooperative kinetics of the constituent enzymes of the complex are indicative of its allosteric properties. Isolation of the alpha-ketoglutarate dehydrogenase complex and its lipoyl dehydrogenase and alpha-ketoglutarate dehydrogenase components in a desensitized state confirms their allosteric nature. It is assumed that NADH effects of isolated alpha-ketoglutarate dehydrogenase is due to a shift in the equilibrium between different oligomeric forms of the enzyme.
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PMID:[Regulation of alpha-ketoglutarate dehydrogenase complex from pigeon breast muscle]. 22 76

Mutants of Escherichia coli K12 with deletions in the nadC-lpd region of the chromosome were obtained for use in studies on the expression of the ace (pyruvate dehydrogenase complex, specific components) and lpd (lipomide dehydrogenase) genes. These were isolated by selecting spontaneous aroP mutants (lacking the general aromatic amino-acid permease and thus resistant to inhibitory aromatic amino-acid analogues) and screening for auxotrophy due to deletions extending into neighbouring genes. From 2892 isolates tested, the AroP- phenotypes of 2322 were confirmed and, of these, 28 stable and independently-derived auxotrophos were designated as deletion mutants. Six nutritionally-distinct categories were recognized: Nad- (8 strains); Nad-Ace-(7): Nad-'Ace-' (3); Ace- (8); 'Ace-' (I); Lpd-(I). The Ace- phenotypes of four isolates designated 'Ace-' were leaky and enzymological studies confirmed that they had less than 7% of parental pyruvate dehydrogenase complex activity. Enzymological studies showed that the 15 Ace- or Nad-Ace- strains all lacked the pyruvate dehydrogenase complex and pyruvate dehydrogenase (EIp) activities and only three retained detectable dihydrolipoamide acetyltransferase (E2p). The one Lpd- strain lacked pyruvate dehydrogenase, dihydrolipoamide acetyltransferase and lipoamide dehydrogenase (E3) activities as well as the activities of the pyruvate and alpha-ketoglutarate dehydrogenase complexes. The results confirmed the gene order nadC-aroP-aceE-aceF-lpd and indicated that no other essential functions are determined by genes within the nadC-lpd region. Resistance to lactate during growth of pps mutants on acetate was directly related to the specific activity of the pyruvate dehydrogenase complex. None of the deletions promoted the high degree of resistance characteristically associated with constitutive expression of the dehydrogenase complex. Six pps mutants having Ace+ or 'Ace-' phenotypes were more sensitive than the parental strains and expression of their ace operons appeared to be affected; most sensitive were the Ace- strains which lacked pyruvate dehydrogenase complex and phosphoenolpyruvate synthetase activities. The lipoamide dehydrogenase activities of the deletion strains (Lpd+) ranged between 30% and 100% of parental levels indicating that expression of their ace operons appeared to be affected; most sensitive were the Ace- strains which lacked pyruvate dehydrogenase complex and phosphoenolpyruvate synthetase activities. The lipoamide dehydrogenase activities of the deletion strains (Lpd+) ranged between 30% and 100% of parental levels indicating that expression of the lpd gene may be affected by the ace operon but can be independent.
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PMID:Biochemical genetics of the alpha-keto acid dehydrogenase complexes of Escherichia coli K12: isolation and biochemical properties of deletion mutants. 32 21

Twenty-eight spontaneous auxotrophic aroP mutants with deletions in the azi--nadC--aroP--aceE--aceF--lpd region of the Escherichia coli K12 chromosome were characterized genetically with respect to various azi, nadC, ace and lpd markers by P1-mediated transduction. One mutant (Kdelta18; aroP--lpddelta) had a deletion which extended through the aceE and aceF genes to end within the lpd gene. The polarity of the ace operon (aceE to aceF) was confirmed. It was concluded that 10 out of 15 deletions generating a strict requirement for acetate terminated in the aceE gene. Of the ten, three mutants (Kdelta22, Cdelta41 and Cdelta41) synthesized detectable dihydrolipoamide acetyltransferase (the aceF gene product) and seven were assumed to possess deletions generating polar effects on aceF gene expression. Five deletions appeared to extend into the aceF gene. A further five deletions, which limited the expression of the ace operon without generating an Ace- phenotype or a complete Ace- phenotype, ended closest to the aroP-proximal aceE markers. The opposite ends of all these deletions appeared to terminate before (10), within (2) or extend beyond (9) the nadC gene. There was no obvious correlation between the deletion end-points and the corresponding lipoamide dehydrogenase activities, which ranged from 30 to 95% of parental levels in different deletion strains. The remaining seven deletions simply extended between the aroP and nadC genes (nad--aroPdelta) without affecting expression of the ace operon. Regulation of the synthesis of the pyruvate and alpha-ketoglutarate dehydrogenase complexes was investigated in some of the parental and deletion strains under different physiological conditions including thiamin-deprivation. The results indicate that the syntheses of the two dehydrogenase complexes are independently regulated. Expression of the lpd gene appears to be coupled to complex synthesis but can be dissociated under some conditions. Mechanisms for regulating lpd gene expression are discussed and an autogenous mechanism involving uncomplexed lipoamide dehydrogenase functioning as a negatively acting repressor at the operator site of an independent lpd gene is proposed as the simplest mechanism which is consistent with all available information.
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PMID:Biochemical genetics of the alpha-keto acid dehydrogenase complexes of Escherichia coli K12: genetic characterization and regulatory properties of deletion mutants. 34 14

The enzymatic defects in a number of Bacillus subtilis mutants of the alpha-ketoglutarate dehydrogenase complex lacking activity have been investigated. Mutants in the citK locus, as well as a series of deletions of unknown length covering the citK locus, are deficient in E1 of the complex, alpha-ketoglutarate dehydrogenase, but have normal activities of E2, dehydrolipoyl transsuccinylase, and E3, lipoamide dehydrogenase. The citK mutants and the citL22 mutant show in vitro complementation of alpha-ketoglutarate dehydrogenase complex activity. The citL22 mutant is severely deficient in lipoamide dehydrogenase activity, and, as a result, lacks activity for both the alpha-ketoglutarate and the pyruvate dehydrogenase complexes. Thus, the E3 components of both complexes are identical. The citL22 mutation maps between ura and metC on the chromosome.
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PMID:Genetics of the alpha-ketoglutarate dehydrogenase complex of Bacillus subtilis. 41 34

The mitochondrial matrix subfractions from rat liver, kidney cortex, brain, heart, and skeletal muscle were isolated and their protein components were resolved by two-dimensional polyacrylamide gel electrophoresis, revealing between 120 and 150 components for each matrix subfraction. Excellent resolution was obtained utilizing a pH 5 to 8 gradient in the first dimension and in 8 to 13% exponential acrylamide gradient in the second dimension, increasing the number of mitochondrial matrix proteins observed 3-fold over one-dimensional systems. Protein components tentatively identified by co-migration with pure enzymes and by known tissue distributions are carbamoyl-phosphate synthetase (EC 2.7.2.5), ornithine transcarbamylase (EC 2.1.3.3), glutamate dehydrogenase (EC 1.4.1.3), pyruvate carboxylase (EC 6.4.1.1), citrate synthase (EC 4.1.3.7), fumarase (EC 4.2.1.2), aconitase (EC 4.2.1.3), alpha-ketoglutarate dehydrogenase (EC 1.2.4.2), dihydrolipoyl transsuccinylase (EC 2.3.1.12), lipoamide dehydrogenase (EC 1.6.4.3), glutamate-aspartate aminotransferase (EC 2.6.1.1), and the two subunits of pyruvate dehydrogenase (EC 1.2.4.1). Protein components unambiguously identified by peptide mapping are citrate synthase, aconitase, and pyruvate carboxylase. The inner membrane subfraction from rat liver mitochondria was also resolved two dimensionally; the alpha and beta subunits of ATPase (F1) (EC 3.6.1.3) were identified by peptide mapping.
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PMID:Resolution of rat mitochondrial matrix proteins by two-dimensional polyacrylamide gel electrophoresis. 44 63

In a case of dihydrolipoyl dehydrogenase deficiency, there was not only an elevation of lactate and alpha-ketoglutarate but also of branched chain amino acids. The levels of branched-chain amino acids varied from the normal range to three times the upper limit of normal during the patient's lifetime, and alloisoleucine was detectable at all times. Examination of postmortem tissues revealed that the activity of branched-chain keto acid dehydrogenases was between zero and 10% of that in control tissues. It is suggested that the multiple defects seen in oxidative decarboxylation in this patient is the consequence of a single genetic deletion of an enzyme common to pyruvate dehydrogenase, alpha-ketoglutarate dehydrogenase, and branched chain keto acid dehydrogenases.
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PMID:A defect in branched-chain amino acid metabolism in a patient with congenital lactic acidosis due to dihydrolipoyl dehydrogenase deficiency. 64 78

An acetate-requiring leaky mutant was induced from Bacillus subtilis 168, and activities of its three alpha-keto acid dehydrogenases were compared with the respectives activities of the parent strain. Both pyruvate and alpha-ketoisovalerate dehydrogenase activities in the mutant were consideralby lower, being only 10-17% of those of the parent, but alpha-ketoglutarate dehydrogenase activity was unchanged. These dehydrogenases are complexed composed of three enzymes: a carboxylase, a lipoic reductase-transacylase, and a dihydrolipoyl dehydrogenase. The carboxylase activity of the affected complexes was no different. Total dihydrolipoyl dehydrogenase activity was only one-third. Thus dihydrolipoyl dehydrogenase is the defective enzyme in the two dehydrogenase complexes; the activity remaining in the mutant is accounted for by the activity of the intact alpha-ketoglutarate dehydrogenase.
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PMID:Activities of alpha-ketoisovalerate, pyruvate, and alpha-ketoglutarate dehydrogenases in a mutant of Bacillus subtilis. 81 42

The redox state of two SH-groups per enzyme subunit has been shown to control the cooperative properties of alpha-ketoglutarate dehydrogenase. These thiols oxidized, alpha-ketoglutarate dehydrogenase does not exhibit any cooperative properties. The enzyme reduction leads to subunit interactions. It has been found that the most effective agent reducing the alpha-ketoglutarate dehydrogenase thiols essential for the cooperativity is dihydrolipoate, one of the intermediates of the overall alpha-ketoglutarate dehydrogenase reaction. The possibility of changing the properties of alpha-ketoglutarate dehydrogenase in the multienzyme complex under the conditions when the lipoic acid integrated into the complex is reduced, has been investigated. Thus, incubation of the alpha-ketoglutarate dehydrogenase complex with NADH has been found to induce the conversion from the non-cooperative form to the cooperative one, presumably through the reduction of lipoic acid bound to the complex in the reaction catalyzed by lipoyl dehydrogenase, the third component of the complex.
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PMID:[Regulation of cooperative properties of alpha-ketoglutarate dehydrogenase by means of thiol-disulfide metabolism]. 191 72

The assembly of alpha-ketoglutarate dehydrogenase complex (KGDC) has been studied in wild-type Saccharomyces cerevisiae and in respiratory-deficient strains (pet) with mutations in KGD1 and KGD2, the structural genes for alpha-ketoglutarate dehydrogenase (KE1) and dihydrolipoyl transsuccinylase (KE2) components, respectively. Mutants unable to express KE1 or KE2 form partial complexes similar to those reported in earlier studies on the resolution and reconstitution of bacterial and mammalian KGDC. Thus mutants lacking KE1 assemble a high-molecular-weight subcomplex consisting of a KE2 core particle with bound dihydrolipoyl dehydrogenase (E3). Similarly, mitochondrial extracts of mutants lacking KE2 contain dimeric KE1 and E3. These components, however, are not associated with each other. The partial complexes detected in the mutants are capable of reconstituting normal KGDC when supplied with the missing subunit. Complete restoration of overall alpha-ketoglutarate dehydrogenase activity is achieved by mixing appropriate ratios of mitochondrial extracts from mutants deficient in KE1 and KE2. The reconstitution of enzymatic activity correlates with binding of KE1 to the KE2-E3 particle to form a complex with the same sedimentation properties as wild-type KGDC. Overexpression of KE2 relative to KE1 results in a preponderance of incompletely assembled complexes with substoichiometric contents of KE1. Formation of a complex with a full complement of KE1 therefore depends on a balanced output of KE1 and KE2 from their respective genes. Biochemical screens of a pet mutant collection have led to the identification of a new gene required for the expression of enzymatically active KGDC. Mitochondria of the mutant have all of the catalytic subunits of KGDC. Sedimentation analysis of these components indicates that while the mutant has a stable KE2-E3 subcomplex, the interaction of KE1 with KE2 core is much weaker in the mutant than in the wild type. The gene product responsible for this phenotype, therefore, appears to function at a late stage of assembly of KGDC, most likely by posttranslational modification of one of the subunits.
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PMID:In vivo assembly of yeast mitochondrial alpha-ketoglutarate dehydrogenase complex. 207

Rat liver lipoamide dehydrogenase (LipDH) was separated into three types on DE-32 column chromatography, but no difference was observed among them in either immunological reactivity or enzymatic properties. A reconstitution experiment of branched-chain alpha-keto acid dehydrogenase complex (BCKADH) revealed that the most anionic type of LipDH was the most effective for the enzyme complex while the three types of LipDH were the same in the affinity for BCKADH subcomplex. All three types of LipDH were equally effective in reconstituting pyruvate dehydrogenase complex, alpha-ketoglutarate dehydrogenase complex and the glycine cleavage system. However, either pyruvate dehydrogenase or alpha-keto-glutarate dehydrogenase complex appeared to involve a certain LipDH in vivo which was firmly integrated into and hardly dissociable from the complex. A broad specificity of LipDH was observed for the glycine cleavage system. When BCKADH reconstitution experiments were carried out with both LipDHs from various sources and purified rat liver BCKADH subcomplex, the effectiveness of animal LipDHs was proportional to the extent of their immunological reactivity to the anti-rat LipDH antibody. However, BCKADH activity was also restored by a certain bacterial LipDH which had no cross-reactivity with the antibody, and LipDHs from some bacterial species, which reacted well with the antibody, showed no effect for the reconstitution of BCKADH. Thus, the determinant(s) of LipDH for the integration into alpha-keto acid dehydrogenase complexes including BCKADH can be its tertiary and/or quarternary structure rather than its primary and secondary structures.
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PMID:Specificity of lipoamide dehydrogenase for alpha-keto acid dehydrogenase complexes and the glycine cleavage system. 213 Dec 88


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