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PubMed1995

[Preparation of monoclonal antibodies to E1-component and their use for studying the pyruvate dehydrogenase complex].

A monoclonal antibody against the E1-component of pigeon breast muscle has been prepared. The dissociation constant of the E1-mAb F7F10 complex was determined to be equal to 5.93.10(-8) M. The cross-reaction between mAb F7F10 and the E1-component of the pyruvate dehydrogenase complex from various species (including human beings) was established. The F7F10 antibody was shown to interact with both alpha- and beta-subunits of E1, which suggests that the amino acid residues in the both subunits are constituents of the antigenic determinant. Binding of the F7F10 antibody to the antigen had no effect on the enzymatic activity of E1 but induced rapid inactivation of the pyruvate dehydrogenase complex in the pyruvate:NAD oxidoreductase reaction. The competition between the F7F10 antibody and the E2-component of the pyruvate dehydrogenase complex for the binding to E1 was revealed by immunoenzymatic analysis. It was concluded that the antigenic determinant and the E1 site responsible for the E1-E2 interaction within the pyruvate dehydrogenase complex may overlap.

Animals↗

PubMed2001

The remarkable structural and functional organization of the eukaryotic pyruvate dehydrogenase complexes.

The three-dimensional reconstruction of the bovine kidney pyruvate dehydrogenase complex (M(r) approximately 7.8 x 10(6)) comprising about 22 molecules of pyruvate dehydrogenase (E(1)) and about 6 molecules of dihydrolipoamide dehydrogenase (E(3)) with its binding protein associated with the 60-subunit dihydrolipoamide acetyltransferase (E(2)) core provides considerable insight into the structural and functional organization of the largest multienzyme complex known. The structure shows that potentially 60 centers for acetyl-CoA synthesis are organized in sets of three at each of the 20 vertices of the pentagonal dodecahedral core. These centers consist of three E(1) molecules bound to one E(2) trimer adjacent to an E(3) molecule in each of 12 pentagonal openings. The E(1) components are anchored to the E(1)-binding domain of the E(2) subunits through an approximately 50-A-long linker. Three of these linkers emanate from the outside edges of the triangular base of the E(2) trimer and form a cage around its base that may shelter the lipoyl domains and the E(1) and E(2) active sites. The docking of the atomic structures of E(1) and the E(1) binding and lipoyl domains of E(2) in the electron microscopy map gives a good fit and indicates that the E(1) active site is approximately 95 A above the base of the trimer. We propose that the lipoyl domains and its tether (swinging arm) rotate about the E(1)-binding domain of E(2,) which is centrally located 45-50 A from the E(1), E(2), and E(3) active sites, and that the highly flexible breathing core augments the transfer of intermediates between active sites.

Animals↗

PubMed1991

Deficiency of the pyruvate dehydrogenase complex and of mitochondrial fatty acid oxidation.

Defects of the pyruvate dehydrogenase complex and of mitochondrial fatty acid oxidation are important causes of disease. Defects of pyruvate dehydrogenase may present in early childhood with severe CNS changes or, as lactic acidosis or later with ataxia. Defects of fatty acid oxidation may present with hypoglycaemic coma, myopathy, liver disease with encephalopathy, cardiomyopathy or sudden infant death. The investigation of both these groups of disorders is difficult and depends upon a combination of biochemical and molecular biology techniques.

Fatty Acids↗

PubMed1981

Lectins activate lymphocyte pyruvate dehydrogenase by a mechanism sensitive to protease inhibitors.

The mitogenic lectins concanavalin A and phytohemagglutinin were found to stimulate pyruvate oxidation in rat mesenteric lymphocytes. Marked cell agglutination accompanied this response. Wheat germ agglutinin, a nonmitogenic lectin, also aggregated lymphocytes but did not cause alteration of pyruvate oxidation. Cell lysates from lectin-treated cells retained their ability to oxidize pyruvate at an elevated rate, indicating that the observed stimulation of pyruvate oxidation was not due to increased transport of labeled pyruvate into the cells. Pyruvate oxidation activity in such lysates was readily sedimented in a mitochondria-enriched cellular fraction, indicating that it reflects mitochondrial pyruvate dehydrogenase. Stimulation of this activity by lectins in intact lymphocytes was inhibited when the cells were incubated under conditions expected to inhibit trypsin-like proteases. Thus, esters of arginine, but not of alanine or tyrosine, blocked stimulation of pyruvate dehydrogenase by the lectins. The data indicate that pyruvate dehydrogenase is activated in lymphocytes treated with mitogenic lectins by a mechanism involving one or more proteolytic reactions. The similarity between the results presented here and those recently reported for insulin action on its target cells [Seals, J. R. & Czech, M. P. (1980) J. Biol. Chem. 255, 6529-6531] suggests that these systems may have similar modes of transmembrane signalling.

Animals↗

PubMed2003

Control of cardiac pyruvate dehydrogenase activity in peroxisome proliferator-activated receptor-alpha transgenic mice.

The pyruvate dehydrogenase enzyme complex (PDC) is rate limiting for glucose oxidation in the heart. Inhibition of PDC by end-product feedback and phosphorylation by pyruvate dehydrogenase kinase (PDK) operate in concert to inhibit PDC activity. Because the transcriptional regulator peroxisome proliferator-activated receptor (PPAR)-alpha increases PDK expression in some tissues, we examined what role PPAR-alpha has in regulating glucose oxidation in hearts from mice overexpressing PPAR-alpha (MHC-PPAR-alpha mice). Glucose oxidation rates were decreased in isolated working hearts from MHC-PPAR-alpha mice compared with wild-type littermates (428 +/- 113 vs. 771 +/- 63 nmol x g dry weight-1x min-1, respectively), which was accompanied by a parallel increase in fatty acid oxidation. However, there was no difference in PDC activity between MHC-PPAR-alpha and wild-type animals, even though the expression of the PDK isoform PDK1 was increased in MHC-PPAR-alpha mice. Glucose oxidation rates in both MHC-PPAR-alpha and wild-type mouse hearts were decreased after 48-h fasting (which increases PPAR-alpha expression) or by treatment of mice with the PPAR-alpha agonist WY-14,643 for 1 wk. Despite this, PDC activity in both animal groups was not altered. Taken together, these data suggest that glucose oxidation rates in the heart can be dramatically altered independent of PDK phosphorylation and inhibition of PDC by PDK. It also suggests that PPAR-alpha activation decreases glucose oxidation in hearts mainly by decreasing the flux of pyruvate through PDC due to negative feedback of PDC by fatty acid oxidation reaction products rather than by the phosphorylated state of the PDC complex.

Animals↗

PubMed1986

Pyruvate dehydrogenase and the path of lactate degradation in Desulfovibrio vulgaris Miyazaki F.

Pyruvate dehydrogenase from Desulfovibrio vulgaris Miyazaki F was partially purified from the soluble fraction of the bacterial sonicate, and characterized. The enzyme catalyzes oxidative decarboxylation of pyruvate to produce acetyl-CoA, in contrast to statements in current review articles in which acetyl phosphate is indicated to be a direct decomposition product of pyruvate in sulfate-reducing bacteria. The established reaction stoichiometry is: pyruvate + CoA + FMN----acetyl-CoA + CO2 + FMNH2. The Km values are 2.9 mM for pyruvate, 32 microM for CoA and 6.7 mumol for FMN. Participation of thiamine diphosphate in the enzymic process was not proven. 2-Oxobutyrate, but not 2-oxoglutarate, can substitute for pyruvate. The three flavin compounds, FMN, FAD, and flavodoxin, as well as clostridial ferredoxin, serve as electron carriers for the enzyme. Thus the enzyme is a kind of pyruvate synthase [EC 1.2.7.1], but acts in the direction of pyruvate degradation in the growing cells. The rate of cytochrome C3 reduction is extremely low, but in the presence of flavodoxin as an electron mediator, the reduction rate of cytochrome C3 becomes faster than the reduction rate of flavodoxin alone. It seems that the physiological electron acceptor for this enzyme is flavodoxin, which might be complexed with cytochrome C3 to produce a very efficient electron transfer system in the cell. The soluble fraction of D. vulgaris cells has been proved to contain, in addition to the pyruvate dehydrogenase, lactate dehydrogenase (Ogata, M., Arihara, K., & Yagi, T. (1981) J. Biochem. 89, 1423-1431), phosphate acetyltransferase and acetate kinase, i.e., all the enzymes necessary to convert lactate to acetate, producing ATP by substrate level phosphorylation.

Acetyl Coenzyme A↗

PubMed1979

Subunit structure of dihydrolipoyl transacetylase component of pyruvate dehydrogenase complex from Escherichia coli.

Limited tryptic digestion of the pyruvate dehydrogenase complex of Escherichia coli or its dihydrolipoyl transacetylase core cleaves the trypsin-sensitive transacetylase subunits into two large fragments, A (lipoyl domain) and D (subunit binding domain). Release of fragments A from the complex does not significantly affect its sedimentation coefficient or its appearance in the electron microscope. Fragment A contains the lipoyl moieties ((3)H-labeled), is acidic with an apparent isoelectric point of about 4.0, has a M(r) of 31,600 as determined by sedimentation equilibrium analysis, and has a swollen or extended structure (f/f(o) = 1.78). Fragment A exhibits anomalous properties, probably due to its acidic nature. It is resistant to staining with Coomassie blue and it migrates on sodium dodecyl sulfate/polyacrylamide gels as if it had a M(r) of 46,000-48,000. Further tryptic digestion converts fragment A into a lipoyl-containing fragment of M(r) 20,000 (fragment B) and eventually into an apparently stable product of estimated M(r) about 10,000 (fragment C). Fragment D has a compact structure of M(r) about 29,600 as determined by sedimentation equilibrium analysis in 6 M guanidinium chloride, and it possesses the intersubunit binding sites of the transacetylase, the binding sites for pyruvate dehydrogenase and dihydrolipoyl dehydrogenase, and the catalytic site for transacetylation. The assemblage of fragments D is responsible for the cube-like appearance of the transacetylase in the electron microscope. High-resolution electron micrographs of the transacetylase show fiber-like extensions, apparently corresponding to tryptic fragment A, surrounding the cube-like core.

Acetyltransferases↗

PubMed1984

Pyruvate dehydrogenase subcomplex with lipoamide dehydrogenase deficiency in a patient with lactic acidosis and branched chain ketoaciduria.

The overall and three component activities of pyruvate dehydrogenase complex were measured in the liver and muscle from a patient who died at 1.9 years with increased concentrations of serum lactate, alpha-ketoglutarate and branched chain amino acids. The component activities of both lipoate acetyltransferase and pyruvate dehydrogenase were similar to those of normal controls, but the overall pyruvate dehydrogenase complex activity was 11 to 30% of controls and lipoamide dehydrogenase activity was not detected. The overall activity was significantly increased by the addition of lipoamide dehydrogenase purified from human liver. Immunochemical studies carried out with antibody prepared against lipoamide dehydrogenase from rat liver, could detect no immunoreactive material in liver and muscle homogenates from the patient, suggesting that the deficiency of lipoamide dehydrogenase activity was due to the lack of enzyme protein.

Acetyltransferases↗

PubMed1984

[Inhibition of kinase activity of the adrenal pyruvate dehydrogenase complex by the mitochondrial component].

A component inhibiting the phosphorylation-linked inactivation of the adrenal pyruvate dehydrogenase complex in the presence of ATP was revealed during purification of the complex from bovine adrenal mitochondria. The degree of the kinase activity inhibition is greater at lower concentrations of ATP. It was assumed that the mitochondrial component screens the kinase active site or the phosphorylation sites of pyruvate dehydrogenase, thus limiting the ATP access to them. Proteins and lipids are incorporated into the component at a ratio 2:1, which is suggestive of its lipoprotein nature. The effect of the mitochondrial component on the kinase activity of the pyruvate dehydrogenase complex is somewhat specific and is unaffected by bovine serum albumin or blood serum lipoproteins.

Adenosine Triphosphate↗

PubMedDate not supplied

[Role of muscle pyruvate dehydrogenase histidine residues in thiamine pyrophosphate binding].

The interaction of diethylpyrocarbonate (DEP) with the pyruvate dehydrogenase component (PDH) isolated from the pyruvate dehydrogenase complex (EC 1.2.4.1) results in a modification of 3-5 histidine residues per mole of enzyme, which simultaneously decreases the enzyme activity. After PDH inhibilion by DEP in the presence of dithiothreitol almost complete reactivation (94%) under the effect of neutral hydroxylamine is observed. In the absence of SH-groups protection incomplete reactivation by hydroxylamine (79%) is found. In the latter case titration with 5,5-dithio--bis-(2-nitrobenzoic acid) in 8 M urea showed that the DEP-modified protein contains less quantity of SH groups (by 4-8) as compared to the native enzyme. It is assumed that the DEP-modified SH-groups are not responsible for the enzyme activity. The differential spectrum of the modified and native PDH showed no changes within the range of 260-300 nm. TPP in combination with Mg2+ (10(-3) M) protectes PDH from being inactivated by DEP. TPP (10(-2) M) reactivates PDH by 70% after its complete inhibition by DEP. Similar protective action is manifested by ATP, ADP and inorganic pyrophosphate in the presence of Mg2+. A kinetic study showed a competitive type of PDH inhibition by DEP with respect to TPP. it is concluded that the histidine residues of PDH are involved in TPP binding.

Catalysis↗

PubMed1978

The effect of fatty acids on the regulation of pyruvate dehydrogenase in perfused rat liver.

The effect of fatty acids on the rate of pyruvate decarboxylation was studied in perfused livers from fed rats. The production of 14CO2 from infused [1-14C]pyruvate was employed as a monitor of the flux through the pyruvate dehydrogenase reaction. A correction for other decarboxylation reactions was made using kinetic analyses. Fatty acid (octanoate or oleate) infusion caused a stimulation of pyruvate decarboxylation at pyruvate concentrations in the perfusate below 1 mM (up to 3-fold at 0.05 mM pyruvate) but decreased the rate to one-third of control rates at pyruvate concentrations near 5 mM. These effects were half-maximal at fatty acid concentrations below 0.1 mM. Infusion of 3-hydroxybutyrate also caused a marked stimulation of pyruvate decarboxylation at low pyruvate concentrations. The data suggest that the mechanism by which fatty acids stimulate the flux through the pyruvate dehydrogenase reaction in perfused liver at low (limiting) pyruvate concentrations involves an acceleration of pyruvate transport into the mitochondrial compartment due to an exchange with acetoacetate. Furthermore, it is proposed that a relationship exists between ketogenesis and the regulation of pyruvate oxidation at pyruvate concentrations approximating conditions in vivo.

Acetoacetates↗

PubMed2001

Inhibition of the Escherichia coli pyruvate dehydrogenase complex E1 subunit and its tyrosine 177 variants by thiamin 2-thiazolone and thiamin 2-thiothiazolone diphosphates. Evidence for reversible tight-binding inhibition.

Variants of the pyruvate dehydrogenase subunit (E1; EC ) of the Escherichia coli pyruvate dehydrogenase multienzyme complex with Y177A and Y177F substitutions were created. Both variants displayed pyruvate dehydrogenase multienzyme complex activity at levels of 11% (Y177A E1) and 7% (Y177F E1) of the parental enzyme. The K(m) values for thiamin diphosphate (ThDP) were 1.58 microm (parental E1) and 6.65 microm (Y177A E1), whereas the Y177F E1 variant was not saturated at 200 microm. According to fluorescence studies, binding of ThDP was unaffected by the Tyr(177) substitutions. The ThDP analogs thiamin 2-thiazolone diphosphate (ThTDP) and thiamin 2-thiothiazolone diphosphate (ThTTDP) behaved as tight-binding inhibitors of parental E1 (K(i) = 0.003 microm for ThTDP and K(i) = 0.064 microm for ThTTDP) and the Y177A and Y177F variants. This analysis revealed that ThTDP and ThTTDP bound to parental E1 via a two-step mechanism, but that ThTDP bound to the Y177A variant via a one-step mechanism. Binding of ThTDP was affected and that of ThTTDP was unaffected by substitutions at Tyr(177). Addition of ThDP or ThTDP to parental E1 resulted in similar CD spectral changes in the near-UV region. In contrast, binding of ThTTDP to either parental E1 or the Y177A and Y177F variants was accompanied by the appearance of a positive band at 330 nm, indicating that ThTTDP was bound in a chiral environment. In combination with x-ray structural evidence on the location of Tyr(177), the kinetic and spectroscopic data suggest that Tyr(177) has a role in stabilization of some transition state(s) in the reaction pathway, starting with the free enzyme and culminating with the first irreversible step (decarboxylation), as well as in reductive acetylation of the dihydrolipoamide acetyltransferase component.

Amino Acid Sequence↗

PubMed1995

Pretranslational regulation of pyruvate dehydrogenase complex subunits in white adipose tissue during the suckling-weaning transition in the rat.

Total pyruvate dehydrogenase complex activity is low in white adipose tissue during the suckling period and increases markedly at weaning on to a high-carbohydrate diet. This is concomitant with an increase in the E1 alpha, E1 beta and E2 subunit protein concentration and their respective mRNAs, suggesting a pretranslational control of this phenomenon. The most marked change is seen for the E1 alpha subunit (17-fold increase in protein concentration). The changes in pyruvate dehydrogenase complex activity and subunit abundance induced by weaning on to a high-carbohydrate diet are precluded if the animals are weaned on to a high-fat diet, suggesting that the nutritional and/or related hormonal changes rather than a developmental stage are responsible for the observed adipose-tissue pyruvate dehydrogenase complex pattern.

Adipose Tissue↗

PubMed1989

Bovine heart pyruvate dehydrogenase kinase stimulation by monovalent ions.

The effects of monovalent ions on endogenous pyruvate dehydrogenase (PDH) kinase activity in purified bovine heart pyruvate dehydrogenase complex were investigated. Activity of PDH kinase was stimulated 1.9-, 1.95-, 1.65-, and 1.4-fold by 10 mM K+, Rb+, NH+4, and Cs+, respectively, whereas Na+ and Li+ had no effect on PDH kinase activity. The crystal radii of stimulatory ions were in the range of 1.33 to 1.69 A while the crystal radii of nonstimulatory ions were in the range of 0.6 to 0.94 A. Stimulation of PDH kinase by monovalent ions was not pH dependent. Protein dilution studies showed that monovalent ion stimulation was measurable within 10 s after protein addition to PDH kinase assays. Furthermore, stimulation occurred at all protein concentrations tested. At ATP concentrations from 12.5 to 25 microM, K+ and NH+4 stimulation was constant from 0 to 110 and 0 to 30 mM, respectively. At higher ATP concentrations, from 50 to 500 microM, K+ and NH+4 stimulation peaked at approximately 30 and 3 mM, respectively, and thereafter declined as the ion concentration increased. Maximal PDH kinase stimulation by K+ or NH+4 also declined as Na+ was increased from 0 to 120 mM, but at a fixed salt concentration of 120 mM, both K+ and NH+4 stimulated PDH kinase activity. Phosphopeptide analysis demonstrated that K+ and NH+4 stimulated phosphorylation at sites 1 and 2, but that site 3 phosphorylation was relatively constant under all conditions. Thiamin pyrophosphate and 5,5'-dithiobis-(2-nitrobenzoate) blocked monovalent ion stimulation half-maximally at 4 and 6 microM, respectively. However, neither thiamin pyrophosphate nor 5,5'-dithiobis-(2-nitrobenzoate) significantly inhibited PDH kinase activity in the absence of monovalent ions. The results indicate that heart PDH kinase stimulation by monovalent ions does not occur by changing the binding equilibrium between PDH and dihydrolipoyl transacetylase core. Instead, monovalent ions bind and exert their regulatory effects at or near the active site of PDH kinase.

Animals↗

PubMed1978

Mechanism of action of the pyruvate dehydrogenase multienzyme complex from Escherichia coli.

The extent of cooperativity among the polypeptide chain components in the overall reaction catalyzed by the pyruvate dehydrogenase multienzyme complex from Escherichia coli has been studied. Selective inactivation of the pyruvate dehydrogenase component with thiamin thiazolone pyrophosphate demonstrates that no cooperativity between this component and the overall catalytic reaction occurs: the amount of overall complex activity is directly proportional to the fraction of active pyruvate dehydrogenase component. The transacetylase component has two lipoic acid residues on each of its polypeptide chains that can be modified by N-[(3)H]ethylmaleimide in the presence of pyruvate and thiamin pyrophosphate. The kinetics of the loss of overall complex activity due to modification of the lipoyl residues on the transacetylase component by maleimide reagents shows that not all lipoic acids are coupled into the overall catalytic reaction and that acyl-group and electron pair transfer involving two or more lipoic acids per catalytic cycle must occur. Finally, full complex activity is found when only half the normal flavin content is present. The results indicate that extensive communication among lipoic acids in acyl-group and electron pair transfer must exist in the normal catalytic mechanism. These results are consistent with the average distances between catalytic sites measured by energy transfer experiments.

Escherichia coli↗

PubMedDate not supplied

[Comparative evaluation of 2 methods for the determination of pyruvate dehydrogenase activity in tissues in avitaminosis B 1 induced by various methods].

A method for determination of pyruvate dehydrogenase activity in mitochondria and tissue homogenates was developed. The method was based on a spectrophotometric monitoring of p-nitroaniline acetylation under conditions required to ensure correct stoichiometric course of the reaction. In rat tissues absolute amounts of the enzyme activity, determined by the method, were found to be several-fold lower as compared with the values determined by the conventional ferricyanide method. Within 24 hrs after a single administration of hydroxythiamin into rats (400 mg per 1 kg of body weight) the pyruvate dehydrogenase activity was decreased 1.5-fold in heart (as estimated by the reaction of p-nitroaniline acetylation) and did not alter in liver tissue. While if the determinations were carried out by the method of terricyanide reduction the enzyme activity was decreas 7- and 2-fold, respectively, in heart and liver tissue. The data obtained suggest that hydroxythiamin impaired reactions of electron transport in tissues; on the other hand, the data obtained showed that the method for determination of the pyruvate dehydrogenase activity, based on the acetylation reaction, was more specific than the conventional method which involved measuring of ferricyanide reduction.

Animals↗

PubMed1998

Isoenzymes of pyruvate dehydrogenase phosphatase. DNA-derived amino acid sequences, expression, and regulation.

Pyruvate dehydrogenase phosphatase (PDP) is one of the few mammalian phosphatases residing within the mitochondrial matrix space. It is responsible for dephosphorylation and reactivation of the pyruvate dehydrogenase complex (PDC) and, by this means, is intimately involved in the regulation of utilization of carbohydrate fuels in mammals. PDP is a dimeric enzyme consisting of catalytic and regulatory subunits. The catalytic subunit of PDP is a Mg2+-dependent enzyme homologous to the cytosolic phosphatases of the 2C family. In the present study, we isolated two cDNAs encoding for mitochondrial phosphatases. The first cDNA is highly homologous to the previously identified cDNA encoding for the catalytic subunit of PDP (PDP1). The second cDNA encodes a previously unknown catalytic subunit of PDP (PDP2). The new phosphatase, expressed as the recombinant protein in Escherichia coli, shows strict substrate specificity toward PDC and does not use phosphorylated branched chain alpha-ketoacid dehydrogenase as substrate. Like PDP1, PDP2 is a Mg2+-dependent enzyme, but its sensitivity to Mg2+ ions is almost 10-fold lower than that of PDP1. In contrast to PDP1, PDP2 is not regulated by Ca2+ ions. Instead, it is sensitive to the biological polyamine spermine, which, in turn, has no effect on the enzymatic activity of PDP1. Western blot analysis of PDP extracted from mitochondria isolated from liver and skeletal muscle revealed that PDP1 is predominantly expressed in mitochondria from skeletal muscle, whereas PDP2 is much more abundant in the liver rather than muscle mitochondria. Both isoenzymes are expressed in mitochondria from 3T3-L1 adipocytes, but the level of expression of PDP2 is considerably higher. These observations are consistent with previous findings on the enzymatic parameters of PDP in adipose tissue. Thus, our results provide the first evidence that there are at least two isoenzymes of PDP in mammals that are different with respect to tissue distribution and kinetic parameters and, therefore, are likely to be different functionally.

3T3 Cells↗

PubMed2006

Role of pyruvate dehydrogenase kinase isoenzyme 4 (PDHK4) in glucose homoeostasis during starvation.

The PDC (pyruvate dehydrogenase complex) is strongly inhibited by phosphorylation during starvation to conserve substrates for gluconeogenesis. The role of PDHK4 (pyruvate dehydrogenase kinase isoenzyme 4) in regulation of PDC by this mechanism was investigated with PDHK4-/- mice (homozygous PDHK4 knockout mice). Starvation lowers blood glucose more in mice lacking PDHK4 than in wild-type mice. The activity state of PDC (percentage dephosphorylated and active) is greater in kidney, gastrocnemius muscle, diaphragm and heart but not in the liver of starved PDHK4-/- mice. Intermediates of the gluconeogenic pathway are lower in concentration in the liver of starved PDHK4-/- mice, consistent with a lower rate of gluconeogenesis due to a substrate supply limitation. The concentration of gluconeogenic substrates is lower in the blood of starved PDHK4-/- mice, consistent with reduced formation in peripheral tissues. Isolated diaphragms from starved PDHK4-/- mice accumulate less lactate and pyruvate because of a faster rate of pyruvate oxidation and a reduced rate of glycolysis. BCAAs (branched chain amino acids) are higher in the blood in starved PDHK4-/- mice, consistent with lower blood alanine levels and the importance of BCAAs as a source of amino groups for alanine formation. Non-esterified fatty acids are also elevated more in the blood of starved PDHK4-/- mice, consistent with lower rates of fatty acid oxidation due to increased rates of glucose and pyruvate oxidation due to greater PDC activity. Up-regulation of PDHK4 in tissues other than the liver is clearly important during starvation for regulation of PDC activity and glucose homoeostasis.

Animals↗