Discovery molecular structure A Trail of Research on Lipoic Acid, L.J. Reed Lipoic Acid Biosynthe... more Discovery molecular structure A Trail of Research on Lipoic Acid, L.J. Reed Lipoic Acid Biosynthesis, N.M. Nesbitt, R.M. Cicchillo, K.H. Lee, T.L. Grove, and S.J. Booker The Search for Potent alpha-lipoic Acid Derivatives: Chemical and Pharmacological Aspects, M. Ben Yakir, Y. Katzhendler, and S. Sasson Novel Indole Lipoic Acid Derivatives: Synthesis and their Antioxidant Effects, A.S. Gurkan and E. Buyukbingol Metabolic aspects The alpha-keto Acid Dehydrogenase Complexes and Glycine Cleavage System: Their Involvement in Pathways of Carbohydrate, Protein, and Fat Metabolism, R.A. Harris, N.H. Jeoung, M. Joshi, and B. Hwang Pyruvate Dehydrogenase Complex Regulation and Lipoic Acid, L. Korotchkina and M.S. Patel Role of Lipoyl Domains in the Function and Regulation of the Mammalian Pyruvate Dehydrogenase Complex, T.E. Roche, T. Peng, L. Hu, Y. Hiromasa, H. Bao, and X. Gong Inactivation and Inhibition of alpha-ketoglutarate Dehydrogenase: Oxidative Modulation of Lipoic Acid, K.M. Humphries, A.C. Nulton-Persson, and L.I. Szweda Lipoate-Protein Ligase A: Structure and Function, K. Fujiwara, H. Hosaka, A. Nakagawa, and Y. Motokawa A Review of the Stability & Preliminary Evaluation of the Plasma Pharmacokinetics of R-lipoic acid & R-Dihydrolipoic Acid Dosage Forms in Human Plasma from Healthy Volunteers, D.A. Carlson, K.L. Young, S.J. Fischer, D.M. Dye, M. Chau, and H. Ulrich The Pharmacokinetics, Metabolism and Renal Excretion of alpha-lipoic Acid and its Main Metabolites in Humans, J. Teichert and R. Preiss Modulation of Cellular Redox and Metabolic Status by Lipoic Acid, D. Han and L. Packer The "Redoxin" Connection of Lipoic Acid, J.A. Barcena, P. Porras, C.A. Padilla, J. Peinado, J.R. Pedrajas, E. Martinez-Galisteo, and R. Requejo Lipoic Acid as an Inducer of Phase II Detoxification Enzymes through Activation of Nrf2-dependent Gene Expression, K. Peterson Shay, S. Shenvi and T.M. Hagen Clinical aspects Deficiency Disorders of Components of the PDH Complex: E2, BP & E3 Deficiencies, J.M. Cameron, M.C. Maj, and B.H. Robinson The Relationship between Primary Biliary Cirrhosis and Lipoic Acid, C. Selmi, X.S. He, C.L. Bowlus, and M.E. Gershwin Effects of Lipoic Acid on Insulin Action in Animal Models of Insulin Resistance, E.J. Henriksen and S. Jacob Activation of Cytoprotective Signaling Pathways by Alpha-Lipoic Acid, A.K. Kiemer and B. Diesel Selenotrisulfide Derivatives of Alpha Lipoic Acid: Potential Use as a Novel Topical Antioxidant, W.T. Self Alpha-Lipoic Acid: A Potent Mitochondrial Nutrient for Improving Memory Deficit, Oxidative Stress, and Mitochondrial Dysfunction, J. Liu Effects of Alpha-Lipoic Acid on AMP-Activated Kinase in Different Tissues: Therapeutic Implications for the Metabolic Syndrome, E.H. Koh, E. H. Cho, M.-S. Kim, J.-Y. Park, and K.-U. Lee
Mammalian pyruvate dehydrogenase complex (PDC) is regulated by phosphorylation-dephosphorylation ... more Mammalian pyruvate dehydrogenase complex (PDC) is regulated by phosphorylation-dephosphorylation of three serine residues (site 1, Ser264; site 2, Ser271; site3, Ser203) of the alpha subunit of pyruvate dehydrogenase (El). Phosphorylation of E l is carried out by four PDK isoenzymes present in cells with differences in tissue-specific distribution, kinetic parameters and regulation. To
— The oxidation of l‐[U‐14C]leucine and l‐[l‐14C]leucine at varying concentrations from 0.1 to 5m... more — The oxidation of l‐[U‐14C]leucine and l‐[l‐14C]leucine at varying concentrations from 0.1 to 5mM to CO2 and the incorporation into cerebral lipids and proteins by brain slices from 1‐week old rats were markedly stimulated by glucose. Although the addition of S mM‐dl‐3‐hydroxybutyrate had no effect on the metabolism of [U‐14C]leucine by brain slices from suckling rats, the stimulatory effects of glucose on the metabolism of l‐[U‐14C]leucine were markedly reduced in the presence of dl‐3‐hydroxybutyrate. The stimulatory effect of glucose on leucine oxidation was, however, not observed in adult rat brain. Furthermore, the incorporation of leucine‐carbon into cerebral lipids and proteins was also very low in the adult brain. The incorporation of l‐[U‐14C]leucine into cerebral lipids by cortex slices was higher during the first 2 postnatal weeks, which then declined to the adult level. During this time span, the oxidation of l‐[U‐14C]leucine to CO2 remained relatively unchanged. The incorporation in vivo of D‐3‐hydroxy[3‐14C]butyrate into cerebral lipids was markedly decreased by acute hyperleucinemia induced by injecting leucine into 9‐day old rats. In in vitro experiments, 5 mM‐leucine had no effect on the oxidation of [U‐14C]glucose to CO2 or its incorporation into lipids by brain slices from 1‐week old rats. However, 5 mM‐leucine inhibited the oxidation of d‐3‐hydroxy‐[3‐14C]butyrate, [3‐14C]acetoacetate and [1‐14C]acetate to CO2 by brain slices, but their incorporation into cerebral lipids was not affected by leucine. In contrast 2‐oxo‐4‐methylvalerate, a deaminated metabolite of leucine, markedly inhibited both the oxidation to CO2 and the incorporation into lipids of labelled glucose, ketone bodies and acetate by cortex slices from 1‐week old rats. These findings suggest that the reduction in the incorporation in vivo of d‐3‐hydroxy[3‐14C]butyrate into cerebral lipids in rats injected with leucine is most likely caused by 2‐oxo‐4‐methylvalerate formed from leucine. Since the concentrations of leucine and 2‐oxo‐4‐methylvalerate in plasma of untreated patients with maple‐syrup urine disease are markedly elevated, our findings are compatible with the possibility that an alteration in the metabolism of glucose and ketone bodies in the brain may contribute to the pathophysiology of this disease.
We have developed overexpression systems for the human branched-chain aminotransferase isoenzymes... more We have developed overexpression systems for the human branched-chain aminotransferase isoenzymes. The enzymes function as dimers and have substrate specificity comparable with the rat enzymes. The human cytosolic enzyme appears to turn over 2-5 times faster than the mitochondrial enzyme, and there may be anion and cation effects on the kinetics of both enzymes. The two proteins demonstrate similar absorption profiles, and the far UV circular dichroism spectra show that no global structural changes occur when the proteins are converted from the pyridoxal to pyridoxamine form. On the other hand, the near UV circular dichroism spectra suggest differences in the local environment surrounding tyrosines within these proteins. Both enzymes require a reducing environment for maximal activity, but the mitochondrial enzyme can be inhibited by nickel ions in the presence of reducing agents, while the cytosolic enzyme is unaffected. Chemical denaturation profiles of the proteins show that there are differences in structural stability. Titration of-SH groups with 5,5dithiobis(2-nitrobenzoic acid) suggests that no disulfide bonds are present in the mitochondrial enzyme and that at least two disulfide bonds are present in the cytosolic enzyme. Two-SH groups are titrated in the native form of the mitochondrial enzyme, leading to complete inhibition of activity, while only one-SH group is titrated in the cytosolic enzyme with no effect on activity. Although these proteins share 58% identity in primary amino acid sequence, the local environment surrounding the active site appears unique for each isoenzyme. Branched-chain aminotransferases (BCAT) 1 catalyze reversible transamination of the short chain aliphatic branched-chain amino acids, leucine, isoleucine, and valine, to their respective ␣-keto acids. Although transamination of branched-chain amino acids in animals and microorganisms was observed in the 1950s (for a review, see Ref. 1), it was not until 1966 that Ichihara and Koyama (2) and Taylor and Jenkins (3) each reported independently that these reactions were catalyzed by a single enzyme.
C'le v c Iund, Ohio 44 106; u nd ' b Depurtme n ts 0.f' Medicine un d Biochemistry, Ge n era1 Cli... more C'le v c Iund, Ohio 44 106; u nd ' b Depurtme n ts 0.f' Medicine un d Biochemistry, Ge n era1 Clin i c d Research Cr n ter .
The oxidation of glutamate by non-synaptic and synaptic mitochondria from brains of 3-, 12- and 2... more The oxidation of glutamate by non-synaptic and synaptic mitochondria from brains of 3-, 12- and 24-month-old rats was studied. With glutamate plus malate as substrates, non-synaptic mitochondria showed higher respiration rates than synaptic mitochondria in all the three age groups studied. The rate of oxidation of L-[1-14C]glutamate and the activities of NAD-glutamate dehydrogenase and aspartate aminotransferase were also higher in non-synaptic mitochondria compared with synaptic mitochondria in three age groups. With glutamate plus malate as substrates, a significant reduction in state 3 respiration was observed in both mitochondrial populations from 12- and 24-month-old rats compared with 3-month-old animals. Although an age-dependent decrease in the oxidation of L-[1-14C]glutamate was observed in both non-synaptic and synaptic mitochondria from aging rats, the oxidation of [1-14C]-2-oxoglutarate was unaltered in non-synaptic and synaptic mitochondria from senescent rats. The activity of NAD-glutamate dehydrogenase was decreased with age in both mitochondrial populations, whereas aspartate aminotransferase was not altered with age. The results indicate that the oxidation rate of glutamate in rat brain mitochondria is decreased during aging.
American Journal of Physiology-endocrinology and Metabolism, Oct 1, 1995
First-generation (1-) male and female rat pups were either reared artificially on a high-carbohyd... more First-generation (1-) male and female rat pups were either reared artificially on a high-carbohydrate (HC) or a high-fat (HF) formula or nursed by mother (MF) from day 4 and weaned onto a stock diet on day 24. 1-HC rats compared with sex-matched control rats (1-HF and 1-MF) were hyperinsulinemic and mildly obese by day 60. We investigated the effect of maternal hyperinsulinemia on the second generation (2-) by intragroup breeding. The 2-HC male and female rats were hyperinsulinemic on day 45, had significantly increased growth rate from approximately day 60 onward, and became obese as evidenced by increased adipose tissue mass due to hypertrophy on day 100. The lipogenic capacity of liver and adipose tissues was significantly higher in the 2-HC than in control rats. Thus the metabolic changes that occurred in the first-generation rats fed a HC formula during early postnatal life not only persisted into their adult life but were also passed on to the next generation.
Amintr acid sequence ecmp~risan of'8 tl nnd 6 JsubunEta of the a-kcta aeici dchydrngenanc (E,] co... more Amintr acid sequence ecmp~risan of'8 tl nnd 6 JsubunEta of the a-kcta aeici dchydrngenanc (E,] component af thr pyruvutc dehydruacnuac complcr and brenehcd-ehrrin wkc~n ticid dckydroganurc enmglox from multiple xpcies was performed by cemputcr analysis. In addition to 2 prcviaullly reeegnircd rcpianr ot'hamaisgy in the 01 subunit, a 3rd rct$an oP cxtcnrlvc homolapy was identified in E,a, and miry bc one of tht? riru invDlvcd in subunit interaction. S,fl containr 4 rcyionr of extensive homology. Region I contains IO amino nridr that WC Ramolo#juur; ta a I&amino acid stretch in Ekh~rieiritr rdi/i E,. Regianr 1 and 3 hrvc scquencc homoiagicn with ether dehydregcnam~ sugyrrting that rhcro fcgionr may be involved in eatnlywix. Byruvntc dchydrogcnaxc: Pyruvntc dchydrelgenttbc complex: Urrnehcd-chain a&eta acid dchydrogcnusc complex: cl-Keto acid dchydregcnrsc corns plcx; Amino acid sequcncccomparioen
Advances in Experimental Medicine and Biology, 1979
During feasting on a balanced carbohydrate, fat, and protein meal resting metabolic rate, body te... more During feasting on a balanced carbohydrate, fat, and protein meal resting metabolic rate, body temperature and respiratory quotient all increase. The dietary components are utilized to replenish and augment glycogen and fat stores in the body. Excessive carbohydrate is also converted to lipid in the liver and stored along with the excessive lipids of dietary origin as triglycerides in adipose tissue, the major fuel storage depot. Amino acids in excess of those needed for protein synthesis are preferentially catabolized over glucose and fat for energy production. This occurs because there are no significant storage sites for amino acids or proteins, and the accumulation of nitrogenous compounds is ill tolerated. During fasting, adipose tissue, muscle, liver, and kidneys work in concert to supply, to convert, and to conserve fuels for the body. During the brief postabsorptive period, blood fuel homeostasis is maintained primarily by hepatic glycogenolysis and adipose tissue lipolysis. As fasting progresses, muscle proteolysis supplies glycogenic amino acids for heightened hepatic gluconeogenesis for a short period of time. After about three days of starvation, the metabolic profile is set to conserve protein and to supply greater quantities of alternate fuels. In particular, free fatty acids and ketone bodies are utilized to maintain energy needs. The ability of the kidney to conserve ketone bodies prevents the loss of large quantities of these valuable fuels in the urine. This delicate interplay among liver, muscle, kidney, and adipose tissue maintains blood fuel homeostasis and allows humans to survive caloric deprivation for extended periods.
The PDC (pyruvate dehydrogenase complex) plays a central role in the maintenance of glucose homoe... more The PDC (pyruvate dehydrogenase complex) plays a central role in the maintenance of glucose homoeostasis in mammals. The carbon flux through the PDC is meticulously controlled by elaborate mechanisms involving post-translational (short-term) phosphorylation/dephosphorylation and transcriptional (long-term) controls. The former regulatory mechanism involving multiple phosphorylation sites and tissue-specific distribution of the dedicated kinases and phosphatases is not only dependent on the interactions among the catalytic and regulatory components of the complex but also sensitive to the intramitochondrial redox state and metabolite levels as indicators of the energy status. Furthermore, differential transcriptional controls of the regulatory components of PDC further add to the complexity needed for long-term tuning of PDC activity for the maintenance of glucose homoeostasis during normal and disease states.
Proceedings of the National Academy of Sciences of the United States of America, Nov 15, 1992
The bacterium Shewanella frigidimarina can grow anaerobically by utilizing Fe(III) as a respirato... more The bacterium Shewanella frigidimarina can grow anaerobically by utilizing Fe(III) as a respiratory electron acceptor. This results in the synthesis of a number of periplasmic c-type cytochromes, which are absent when the organism is grown in the absence of added Fe(III). One cytochrome, IfcA, is synthesized when Fe(III) is present as the sole respiratory electron acceptor or when it is present in combination with oxygen, fumarate, or nitrate. The ifcA gene was thus selected for a study of iron-responsive gene regulation of respiratory proteins in S. frigidimarina. The monocistronic ifcA gene clusters with two other monocistronic genes, ifcO, encoding a putative outer membrane porin, and ifcR, encoding a putative transcriptional regulator of the LysR superfamily. Analysis of transcription of all three genes under a range of growth conditions in the wild type and an ifcR insertion mutant and analysis of a strain that constitutively expresses ifcR revealed that iron regulation is exerted at the level of ifcR transcription. In the presence of Fe(III) IfcR is synthesized and acts positively to regulate expression of ifcO and ifcA. Control of Fe(III) respiration by this novel regulatory system differs markedly from Fur-mediated regulation of iron assimilation, in which Fur serves as an Fe(II)-activated repressor.
Changes in the uptake of pyruvate by nonsynaptic and synaptic mitochondria from brains of young a... more Changes in the uptake of pyruvate by nonsynaptic and synaptic mitochondria from brains of young adult and old rats were investigated. An age-dependent decrease in State 3 respiration in the presence of pyruvate plus malate as substrate was observed in cerebral mitochondrial populations but not in liver mitochondria. Addition of exogenous cytochrome c to nonsynaptic and synaptic mitochondria enhanced the rate of State 3 respiration but the age-dependent decrease in State 3 respiration persisted in both types of mitochondria. A decrease in the uptake of pyruvate as measured by the inhibitor-stop and rapid centrifugation techniques was observed in both nonsynaptic and synaptic mitochondria from 24-month-old rats compared to 3-month-old rats. The results suggest that the decrease in the uptake of pyruvate may be one of the factors responsible for the observed reduction in State 3 respiration in the presence of pyruvate plus malate by both nonsynaptic and synaptic mitochondria from brains of senescent rats compared to young adults.
Splanchnic exchange rates of glucose, min. There was a positive correlation beacetoacetate, P-hyd... more Splanchnic exchange rates of glucose, min. There was a positive correlation beacetoacetate, P-hydroxybutyrate, lactate, tween free fatty acid uptake and ketonepyruvate, glycerol, alanine, glutamine, body release. Net splanchnic acetoacetate glutamate, free fatty acids, and triglycerrelease was 0.50 (0.05-0.92) mmole/min ides were measured in eight patients and /3-hydroxybutyrate release was 0.35 during moderate to severe diabetic keto-(-0.16 to 0.84) mmole/min. Total ketoneacidosis. Their arterial glucose concentra-body release was 0.85 (0.37-1.61) tion was 20.68 (9.80-52.79) pmole/liter mmole/min. The wide ranges of net and total ketone-body concentration was splanchnic glucose and ketone-body pro-10.38 (6.04-15.81) mmole/liter. Net duction rates show the heterogeneous splanchnic glucose release was 0.77 characteristics of the diabetic patient in (0.09-2.44) mmole/min. Gluconeogenesis ketoacidosis. It is concluded that the hyaccounted for about one-half of net perglycemia and hyperketonemia of splanchnic glucose release, assuming diabetic ketoacidosis is due to the lack of quantitative conversion of net splanchnic reciprocity among rates of hepatic glyextracted lactate, pyruvate, glycerol, cogenolysis, gluconeogenesis, and ketoalanine, and a-ketoglutarate equivalents genesis resulting in inappropriate net to glucose. Net splanchnic free fatty acid splanchnic release of glucose and ketone extraction was 0.24 (0.09-0.52) mmole/ bodies.
Deoxynucleotide sequencing of a cDNA for the dihydrolipoamide acetyltransferase (PDC-E,) componen... more Deoxynucleotide sequencing of a cDNA for the dihydrolipoamide acetyltransferase (PDC-E,) component of human pyruvate dehydrogenase complex (PDC) revealed an open reading frame of 1848 base pairs corresponding to a leader sequence of 54 amino acids and a mature protein of 561 amino acids (59551 Da). Both an amino-terminal lipoyl-bearing domain and a carboxy-terminal catalytic domain are present in the deduced amino acid sequence. The lipoyl-bearing domain contains two repeating units of 127 amino acids, each harboring one lipoic acid-binding lysine. Thus, mammalian PDC-E, differs as to the number of lipoic acid-binding sites from other dihydrolipoamide acyltransferases in both prokaryotic and eukaryotic organisms.
Der Substratstoffwechsel von Organen kann grundsatzlich auf zweierlei Weise studiert werden: Einm... more Der Substratstoffwechsel von Organen kann grundsatzlich auf zweierlei Weise studiert werden: Einmal mit Hilfe der Kathetertechnik und andererseits mit Hilfe der Isotopentechnik. Bei der Kathetertechnik wird die Konzentration des jeweiligen Substrates vor und hinter dem Organ simultan gemessen. Gleichzeitig wird der Durchflus des Organs pro Zeiteinheit bestimmt. Werden diese Messungen unter konstanten Bedingungen durchgefuhrt, kann man nach dem Fickschen Prinzip aus der Konzentrationsdifferenz der Substrate × der Durchblutung die Aufnahme oder Abgabe eines Substrats kalkulieren [1]. Wahrend es nicht schwierig ist, den Substratflus in ein Organ durch Messung der Konzentration der Substrate in einer zuganglichen Arterie zu bestimmen, wird es schwieriger im lebenden Organismus, das aus inneren Organen austretende Blut zu gewinnen. Dies ist praktisch nur moglich mit Hilfe eines Katheters, der im venosen Gefassystem hinter das Organ vorgeschoben wird. Trotzdem fanden bereits am Ende des letzten Jahrhunderts Messungen des Substratumsatzes von Geweben am lebenden Tier statt. Ein gutes Beispiel sind die Studien von Chaveau und Kaufmann aus dem Jahre 1887 [2], die an einem leicht zuganglichen Muskel, dem Lippenmuskel am Pferd, den Einflus der Muskelaktivitat auf den Substratumsatz beobachteten.
Glucose is the major fuel for oxidative metabolism in the developing fetus. A continuous supply o... more Glucose is the major fuel for oxidative metabolism in the developing fetus. A continuous supply of glucose and other nutrients from the maternal circulation to the fetus via the placenta is essential for normal fetal development. The initial metabolism of glucose to pyruvate occurs by the glycolytic pathway; further oxidation of pyruvate to C02 takes place through the tricarboxylic acid cycle in mitochondria. These two pathways are functionally linked by the pyruvate dehydrogenase complex (PDC) in the mitochondria. The level of oxidative metabolism of pyruvate continually increases in different tissues beginning in the early postnatal period, and gradually reaches the maximal levels observed in tissues from adults. In this chapter, we will discuss the structure, function and regulation of mammalian PDC with special emphasis on recent advances in the cloning of several components of the complex. Additionally, the expression of PDC activity in differentiating cultured cells and developing animals will be discussed. The importance of normal pyruvate oxidation during early mammalian development will be underlined by discussing the effects of inborn errors of PDC.
Acta Crystallographica Section D-biological Crystallography, Mar 1, 2001
TehB is an S-adenosyl-l-methionine (SAM) dependent methyltransferase that detoxifies tellurite in... more TehB is an S-adenosyl-l-methionine (SAM) dependent methyltransferase that detoxifies tellurite in bacteria. The Escherichia coli TehB protein was purified and crystallized in the presence of both SAM and sinefungin. The TehB-SAM and TehB-sinefungin crystals both diffracted X-rays to 1.9 Å resolution. The TehB-SAM crystals belonged to space group C2, with unit-cell parameters a = 60.0, b = 56.1, c = 130.6 Å , = 97.9. The TehB-sinefungin crystals belonged to space group P2 1 , with unit-cell parameters a = 59.1, b = 55.5, c = 129.7 Å , = 95.9 .
Discovery molecular structure A Trail of Research on Lipoic Acid, L.J. Reed Lipoic Acid Biosynthe... more Discovery molecular structure A Trail of Research on Lipoic Acid, L.J. Reed Lipoic Acid Biosynthesis, N.M. Nesbitt, R.M. Cicchillo, K.H. Lee, T.L. Grove, and S.J. Booker The Search for Potent alpha-lipoic Acid Derivatives: Chemical and Pharmacological Aspects, M. Ben Yakir, Y. Katzhendler, and S. Sasson Novel Indole Lipoic Acid Derivatives: Synthesis and their Antioxidant Effects, A.S. Gurkan and E. Buyukbingol Metabolic aspects The alpha-keto Acid Dehydrogenase Complexes and Glycine Cleavage System: Their Involvement in Pathways of Carbohydrate, Protein, and Fat Metabolism, R.A. Harris, N.H. Jeoung, M. Joshi, and B. Hwang Pyruvate Dehydrogenase Complex Regulation and Lipoic Acid, L. Korotchkina and M.S. Patel Role of Lipoyl Domains in the Function and Regulation of the Mammalian Pyruvate Dehydrogenase Complex, T.E. Roche, T. Peng, L. Hu, Y. Hiromasa, H. Bao, and X. Gong Inactivation and Inhibition of alpha-ketoglutarate Dehydrogenase: Oxidative Modulation of Lipoic Acid, K.M. Humphries, A.C. Nulton-Persson, and L.I. Szweda Lipoate-Protein Ligase A: Structure and Function, K. Fujiwara, H. Hosaka, A. Nakagawa, and Y. Motokawa A Review of the Stability & Preliminary Evaluation of the Plasma Pharmacokinetics of R-lipoic acid & R-Dihydrolipoic Acid Dosage Forms in Human Plasma from Healthy Volunteers, D.A. Carlson, K.L. Young, S.J. Fischer, D.M. Dye, M. Chau, and H. Ulrich The Pharmacokinetics, Metabolism and Renal Excretion of alpha-lipoic Acid and its Main Metabolites in Humans, J. Teichert and R. Preiss Modulation of Cellular Redox and Metabolic Status by Lipoic Acid, D. Han and L. Packer The "Redoxin" Connection of Lipoic Acid, J.A. Barcena, P. Porras, C.A. Padilla, J. Peinado, J.R. Pedrajas, E. Martinez-Galisteo, and R. Requejo Lipoic Acid as an Inducer of Phase II Detoxification Enzymes through Activation of Nrf2-dependent Gene Expression, K. Peterson Shay, S. Shenvi and T.M. Hagen Clinical aspects Deficiency Disorders of Components of the PDH Complex: E2, BP & E3 Deficiencies, J.M. Cameron, M.C. Maj, and B.H. Robinson The Relationship between Primary Biliary Cirrhosis and Lipoic Acid, C. Selmi, X.S. He, C.L. Bowlus, and M.E. Gershwin Effects of Lipoic Acid on Insulin Action in Animal Models of Insulin Resistance, E.J. Henriksen and S. Jacob Activation of Cytoprotective Signaling Pathways by Alpha-Lipoic Acid, A.K. Kiemer and B. Diesel Selenotrisulfide Derivatives of Alpha Lipoic Acid: Potential Use as a Novel Topical Antioxidant, W.T. Self Alpha-Lipoic Acid: A Potent Mitochondrial Nutrient for Improving Memory Deficit, Oxidative Stress, and Mitochondrial Dysfunction, J. Liu Effects of Alpha-Lipoic Acid on AMP-Activated Kinase in Different Tissues: Therapeutic Implications for the Metabolic Syndrome, E.H. Koh, E. H. Cho, M.-S. Kim, J.-Y. Park, and K.-U. Lee
Mammalian pyruvate dehydrogenase complex (PDC) is regulated by phosphorylation-dephosphorylation ... more Mammalian pyruvate dehydrogenase complex (PDC) is regulated by phosphorylation-dephosphorylation of three serine residues (site 1, Ser264; site 2, Ser271; site3, Ser203) of the alpha subunit of pyruvate dehydrogenase (El). Phosphorylation of E l is carried out by four PDK isoenzymes present in cells with differences in tissue-specific distribution, kinetic parameters and regulation. To
— The oxidation of l‐[U‐14C]leucine and l‐[l‐14C]leucine at varying concentrations from 0.1 to 5m... more — The oxidation of l‐[U‐14C]leucine and l‐[l‐14C]leucine at varying concentrations from 0.1 to 5mM to CO2 and the incorporation into cerebral lipids and proteins by brain slices from 1‐week old rats were markedly stimulated by glucose. Although the addition of S mM‐dl‐3‐hydroxybutyrate had no effect on the metabolism of [U‐14C]leucine by brain slices from suckling rats, the stimulatory effects of glucose on the metabolism of l‐[U‐14C]leucine were markedly reduced in the presence of dl‐3‐hydroxybutyrate. The stimulatory effect of glucose on leucine oxidation was, however, not observed in adult rat brain. Furthermore, the incorporation of leucine‐carbon into cerebral lipids and proteins was also very low in the adult brain. The incorporation of l‐[U‐14C]leucine into cerebral lipids by cortex slices was higher during the first 2 postnatal weeks, which then declined to the adult level. During this time span, the oxidation of l‐[U‐14C]leucine to CO2 remained relatively unchanged. The incorporation in vivo of D‐3‐hydroxy[3‐14C]butyrate into cerebral lipids was markedly decreased by acute hyperleucinemia induced by injecting leucine into 9‐day old rats. In in vitro experiments, 5 mM‐leucine had no effect on the oxidation of [U‐14C]glucose to CO2 or its incorporation into lipids by brain slices from 1‐week old rats. However, 5 mM‐leucine inhibited the oxidation of d‐3‐hydroxy‐[3‐14C]butyrate, [3‐14C]acetoacetate and [1‐14C]acetate to CO2 by brain slices, but their incorporation into cerebral lipids was not affected by leucine. In contrast 2‐oxo‐4‐methylvalerate, a deaminated metabolite of leucine, markedly inhibited both the oxidation to CO2 and the incorporation into lipids of labelled glucose, ketone bodies and acetate by cortex slices from 1‐week old rats. These findings suggest that the reduction in the incorporation in vivo of d‐3‐hydroxy[3‐14C]butyrate into cerebral lipids in rats injected with leucine is most likely caused by 2‐oxo‐4‐methylvalerate formed from leucine. Since the concentrations of leucine and 2‐oxo‐4‐methylvalerate in plasma of untreated patients with maple‐syrup urine disease are markedly elevated, our findings are compatible with the possibility that an alteration in the metabolism of glucose and ketone bodies in the brain may contribute to the pathophysiology of this disease.
We have developed overexpression systems for the human branched-chain aminotransferase isoenzymes... more We have developed overexpression systems for the human branched-chain aminotransferase isoenzymes. The enzymes function as dimers and have substrate specificity comparable with the rat enzymes. The human cytosolic enzyme appears to turn over 2-5 times faster than the mitochondrial enzyme, and there may be anion and cation effects on the kinetics of both enzymes. The two proteins demonstrate similar absorption profiles, and the far UV circular dichroism spectra show that no global structural changes occur when the proteins are converted from the pyridoxal to pyridoxamine form. On the other hand, the near UV circular dichroism spectra suggest differences in the local environment surrounding tyrosines within these proteins. Both enzymes require a reducing environment for maximal activity, but the mitochondrial enzyme can be inhibited by nickel ions in the presence of reducing agents, while the cytosolic enzyme is unaffected. Chemical denaturation profiles of the proteins show that there are differences in structural stability. Titration of-SH groups with 5,5dithiobis(2-nitrobenzoic acid) suggests that no disulfide bonds are present in the mitochondrial enzyme and that at least two disulfide bonds are present in the cytosolic enzyme. Two-SH groups are titrated in the native form of the mitochondrial enzyme, leading to complete inhibition of activity, while only one-SH group is titrated in the cytosolic enzyme with no effect on activity. Although these proteins share 58% identity in primary amino acid sequence, the local environment surrounding the active site appears unique for each isoenzyme. Branched-chain aminotransferases (BCAT) 1 catalyze reversible transamination of the short chain aliphatic branched-chain amino acids, leucine, isoleucine, and valine, to their respective ␣-keto acids. Although transamination of branched-chain amino acids in animals and microorganisms was observed in the 1950s (for a review, see Ref. 1), it was not until 1966 that Ichihara and Koyama (2) and Taylor and Jenkins (3) each reported independently that these reactions were catalyzed by a single enzyme.
C'le v c Iund, Ohio 44 106; u nd ' b Depurtme n ts 0.f' Medicine un d Biochemistry, Ge n era1 Cli... more C'le v c Iund, Ohio 44 106; u nd ' b Depurtme n ts 0.f' Medicine un d Biochemistry, Ge n era1 Clin i c d Research Cr n ter .
The oxidation of glutamate by non-synaptic and synaptic mitochondria from brains of 3-, 12- and 2... more The oxidation of glutamate by non-synaptic and synaptic mitochondria from brains of 3-, 12- and 24-month-old rats was studied. With glutamate plus malate as substrates, non-synaptic mitochondria showed higher respiration rates than synaptic mitochondria in all the three age groups studied. The rate of oxidation of L-[1-14C]glutamate and the activities of NAD-glutamate dehydrogenase and aspartate aminotransferase were also higher in non-synaptic mitochondria compared with synaptic mitochondria in three age groups. With glutamate plus malate as substrates, a significant reduction in state 3 respiration was observed in both mitochondrial populations from 12- and 24-month-old rats compared with 3-month-old animals. Although an age-dependent decrease in the oxidation of L-[1-14C]glutamate was observed in both non-synaptic and synaptic mitochondria from aging rats, the oxidation of [1-14C]-2-oxoglutarate was unaltered in non-synaptic and synaptic mitochondria from senescent rats. The activity of NAD-glutamate dehydrogenase was decreased with age in both mitochondrial populations, whereas aspartate aminotransferase was not altered with age. The results indicate that the oxidation rate of glutamate in rat brain mitochondria is decreased during aging.
American Journal of Physiology-endocrinology and Metabolism, Oct 1, 1995
First-generation (1-) male and female rat pups were either reared artificially on a high-carbohyd... more First-generation (1-) male and female rat pups were either reared artificially on a high-carbohydrate (HC) or a high-fat (HF) formula or nursed by mother (MF) from day 4 and weaned onto a stock diet on day 24. 1-HC rats compared with sex-matched control rats (1-HF and 1-MF) were hyperinsulinemic and mildly obese by day 60. We investigated the effect of maternal hyperinsulinemia on the second generation (2-) by intragroup breeding. The 2-HC male and female rats were hyperinsulinemic on day 45, had significantly increased growth rate from approximately day 60 onward, and became obese as evidenced by increased adipose tissue mass due to hypertrophy on day 100. The lipogenic capacity of liver and adipose tissues was significantly higher in the 2-HC than in control rats. Thus the metabolic changes that occurred in the first-generation rats fed a HC formula during early postnatal life not only persisted into their adult life but were also passed on to the next generation.
Amintr acid sequence ecmp~risan of'8 tl nnd 6 JsubunEta of the a-kcta aeici dchydrngenanc (E,] co... more Amintr acid sequence ecmp~risan of'8 tl nnd 6 JsubunEta of the a-kcta aeici dchydrngenanc (E,] component af thr pyruvutc dehydruacnuac complcr and brenehcd-ehrrin wkc~n ticid dckydroganurc enmglox from multiple xpcies was performed by cemputcr analysis. In addition to 2 prcviaullly reeegnircd rcpianr ot'hamaisgy in the 01 subunit, a 3rd rct$an oP cxtcnrlvc homolapy was identified in E,a, and miry bc one of tht? riru invDlvcd in subunit interaction. S,fl containr 4 rcyionr of extensive homology. Region I contains IO amino nridr that WC Ramolo#juur; ta a I&amino acid stretch in Ekh~rieiritr rdi/i E,. Regianr 1 and 3 hrvc scquencc homoiagicn with ether dehydregcnam~ sugyrrting that rhcro fcgionr may be involved in eatnlywix. Byruvntc dchydrogcnaxc: Pyruvntc dchydrelgenttbc complex: Urrnehcd-chain a&eta acid dchydrogcnusc complex: cl-Keto acid dchydregcnrsc corns plcx; Amino acid sequcncccomparioen
Advances in Experimental Medicine and Biology, 1979
During feasting on a balanced carbohydrate, fat, and protein meal resting metabolic rate, body te... more During feasting on a balanced carbohydrate, fat, and protein meal resting metabolic rate, body temperature and respiratory quotient all increase. The dietary components are utilized to replenish and augment glycogen and fat stores in the body. Excessive carbohydrate is also converted to lipid in the liver and stored along with the excessive lipids of dietary origin as triglycerides in adipose tissue, the major fuel storage depot. Amino acids in excess of those needed for protein synthesis are preferentially catabolized over glucose and fat for energy production. This occurs because there are no significant storage sites for amino acids or proteins, and the accumulation of nitrogenous compounds is ill tolerated. During fasting, adipose tissue, muscle, liver, and kidneys work in concert to supply, to convert, and to conserve fuels for the body. During the brief postabsorptive period, blood fuel homeostasis is maintained primarily by hepatic glycogenolysis and adipose tissue lipolysis. As fasting progresses, muscle proteolysis supplies glycogenic amino acids for heightened hepatic gluconeogenesis for a short period of time. After about three days of starvation, the metabolic profile is set to conserve protein and to supply greater quantities of alternate fuels. In particular, free fatty acids and ketone bodies are utilized to maintain energy needs. The ability of the kidney to conserve ketone bodies prevents the loss of large quantities of these valuable fuels in the urine. This delicate interplay among liver, muscle, kidney, and adipose tissue maintains blood fuel homeostasis and allows humans to survive caloric deprivation for extended periods.
The PDC (pyruvate dehydrogenase complex) plays a central role in the maintenance of glucose homoe... more The PDC (pyruvate dehydrogenase complex) plays a central role in the maintenance of glucose homoeostasis in mammals. The carbon flux through the PDC is meticulously controlled by elaborate mechanisms involving post-translational (short-term) phosphorylation/dephosphorylation and transcriptional (long-term) controls. The former regulatory mechanism involving multiple phosphorylation sites and tissue-specific distribution of the dedicated kinases and phosphatases is not only dependent on the interactions among the catalytic and regulatory components of the complex but also sensitive to the intramitochondrial redox state and metabolite levels as indicators of the energy status. Furthermore, differential transcriptional controls of the regulatory components of PDC further add to the complexity needed for long-term tuning of PDC activity for the maintenance of glucose homoeostasis during normal and disease states.
Proceedings of the National Academy of Sciences of the United States of America, Nov 15, 1992
The bacterium Shewanella frigidimarina can grow anaerobically by utilizing Fe(III) as a respirato... more The bacterium Shewanella frigidimarina can grow anaerobically by utilizing Fe(III) as a respiratory electron acceptor. This results in the synthesis of a number of periplasmic c-type cytochromes, which are absent when the organism is grown in the absence of added Fe(III). One cytochrome, IfcA, is synthesized when Fe(III) is present as the sole respiratory electron acceptor or when it is present in combination with oxygen, fumarate, or nitrate. The ifcA gene was thus selected for a study of iron-responsive gene regulation of respiratory proteins in S. frigidimarina. The monocistronic ifcA gene clusters with two other monocistronic genes, ifcO, encoding a putative outer membrane porin, and ifcR, encoding a putative transcriptional regulator of the LysR superfamily. Analysis of transcription of all three genes under a range of growth conditions in the wild type and an ifcR insertion mutant and analysis of a strain that constitutively expresses ifcR revealed that iron regulation is exerted at the level of ifcR transcription. In the presence of Fe(III) IfcR is synthesized and acts positively to regulate expression of ifcO and ifcA. Control of Fe(III) respiration by this novel regulatory system differs markedly from Fur-mediated regulation of iron assimilation, in which Fur serves as an Fe(II)-activated repressor.
Changes in the uptake of pyruvate by nonsynaptic and synaptic mitochondria from brains of young a... more Changes in the uptake of pyruvate by nonsynaptic and synaptic mitochondria from brains of young adult and old rats were investigated. An age-dependent decrease in State 3 respiration in the presence of pyruvate plus malate as substrate was observed in cerebral mitochondrial populations but not in liver mitochondria. Addition of exogenous cytochrome c to nonsynaptic and synaptic mitochondria enhanced the rate of State 3 respiration but the age-dependent decrease in State 3 respiration persisted in both types of mitochondria. A decrease in the uptake of pyruvate as measured by the inhibitor-stop and rapid centrifugation techniques was observed in both nonsynaptic and synaptic mitochondria from 24-month-old rats compared to 3-month-old rats. The results suggest that the decrease in the uptake of pyruvate may be one of the factors responsible for the observed reduction in State 3 respiration in the presence of pyruvate plus malate by both nonsynaptic and synaptic mitochondria from brains of senescent rats compared to young adults.
Splanchnic exchange rates of glucose, min. There was a positive correlation beacetoacetate, P-hyd... more Splanchnic exchange rates of glucose, min. There was a positive correlation beacetoacetate, P-hydroxybutyrate, lactate, tween free fatty acid uptake and ketonepyruvate, glycerol, alanine, glutamine, body release. Net splanchnic acetoacetate glutamate, free fatty acids, and triglycerrelease was 0.50 (0.05-0.92) mmole/min ides were measured in eight patients and /3-hydroxybutyrate release was 0.35 during moderate to severe diabetic keto-(-0.16 to 0.84) mmole/min. Total ketoneacidosis. Their arterial glucose concentra-body release was 0.85 (0.37-1.61) tion was 20.68 (9.80-52.79) pmole/liter mmole/min. The wide ranges of net and total ketone-body concentration was splanchnic glucose and ketone-body pro-10.38 (6.04-15.81) mmole/liter. Net duction rates show the heterogeneous splanchnic glucose release was 0.77 characteristics of the diabetic patient in (0.09-2.44) mmole/min. Gluconeogenesis ketoacidosis. It is concluded that the hyaccounted for about one-half of net perglycemia and hyperketonemia of splanchnic glucose release, assuming diabetic ketoacidosis is due to the lack of quantitative conversion of net splanchnic reciprocity among rates of hepatic glyextracted lactate, pyruvate, glycerol, cogenolysis, gluconeogenesis, and ketoalanine, and a-ketoglutarate equivalents genesis resulting in inappropriate net to glucose. Net splanchnic free fatty acid splanchnic release of glucose and ketone extraction was 0.24 (0.09-0.52) mmole/ bodies.
Deoxynucleotide sequencing of a cDNA for the dihydrolipoamide acetyltransferase (PDC-E,) componen... more Deoxynucleotide sequencing of a cDNA for the dihydrolipoamide acetyltransferase (PDC-E,) component of human pyruvate dehydrogenase complex (PDC) revealed an open reading frame of 1848 base pairs corresponding to a leader sequence of 54 amino acids and a mature protein of 561 amino acids (59551 Da). Both an amino-terminal lipoyl-bearing domain and a carboxy-terminal catalytic domain are present in the deduced amino acid sequence. The lipoyl-bearing domain contains two repeating units of 127 amino acids, each harboring one lipoic acid-binding lysine. Thus, mammalian PDC-E, differs as to the number of lipoic acid-binding sites from other dihydrolipoamide acyltransferases in both prokaryotic and eukaryotic organisms.
Der Substratstoffwechsel von Organen kann grundsatzlich auf zweierlei Weise studiert werden: Einm... more Der Substratstoffwechsel von Organen kann grundsatzlich auf zweierlei Weise studiert werden: Einmal mit Hilfe der Kathetertechnik und andererseits mit Hilfe der Isotopentechnik. Bei der Kathetertechnik wird die Konzentration des jeweiligen Substrates vor und hinter dem Organ simultan gemessen. Gleichzeitig wird der Durchflus des Organs pro Zeiteinheit bestimmt. Werden diese Messungen unter konstanten Bedingungen durchgefuhrt, kann man nach dem Fickschen Prinzip aus der Konzentrationsdifferenz der Substrate × der Durchblutung die Aufnahme oder Abgabe eines Substrats kalkulieren [1]. Wahrend es nicht schwierig ist, den Substratflus in ein Organ durch Messung der Konzentration der Substrate in einer zuganglichen Arterie zu bestimmen, wird es schwieriger im lebenden Organismus, das aus inneren Organen austretende Blut zu gewinnen. Dies ist praktisch nur moglich mit Hilfe eines Katheters, der im venosen Gefassystem hinter das Organ vorgeschoben wird. Trotzdem fanden bereits am Ende des letzten Jahrhunderts Messungen des Substratumsatzes von Geweben am lebenden Tier statt. Ein gutes Beispiel sind die Studien von Chaveau und Kaufmann aus dem Jahre 1887 [2], die an einem leicht zuganglichen Muskel, dem Lippenmuskel am Pferd, den Einflus der Muskelaktivitat auf den Substratumsatz beobachteten.
Glucose is the major fuel for oxidative metabolism in the developing fetus. A continuous supply o... more Glucose is the major fuel for oxidative metabolism in the developing fetus. A continuous supply of glucose and other nutrients from the maternal circulation to the fetus via the placenta is essential for normal fetal development. The initial metabolism of glucose to pyruvate occurs by the glycolytic pathway; further oxidation of pyruvate to C02 takes place through the tricarboxylic acid cycle in mitochondria. These two pathways are functionally linked by the pyruvate dehydrogenase complex (PDC) in the mitochondria. The level of oxidative metabolism of pyruvate continually increases in different tissues beginning in the early postnatal period, and gradually reaches the maximal levels observed in tissues from adults. In this chapter, we will discuss the structure, function and regulation of mammalian PDC with special emphasis on recent advances in the cloning of several components of the complex. Additionally, the expression of PDC activity in differentiating cultured cells and developing animals will be discussed. The importance of normal pyruvate oxidation during early mammalian development will be underlined by discussing the effects of inborn errors of PDC.
Acta Crystallographica Section D-biological Crystallography, Mar 1, 2001
TehB is an S-adenosyl-l-methionine (SAM) dependent methyltransferase that detoxifies tellurite in... more TehB is an S-adenosyl-l-methionine (SAM) dependent methyltransferase that detoxifies tellurite in bacteria. The Escherichia coli TehB protein was purified and crystallized in the presence of both SAM and sinefungin. The TehB-SAM and TehB-sinefungin crystals both diffracted X-rays to 1.9 Å resolution. The TehB-SAM crystals belonged to space group C2, with unit-cell parameters a = 60.0, b = 56.1, c = 130.6 Å , = 97.9. The TehB-sinefungin crystals belonged to space group P2 1 , with unit-cell parameters a = 59.1, b = 55.5, c = 129.7 Å , = 95.9 .
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Papers by Mulchand Patel