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Sample records for 1-semialdehyde aminotransferase gsa-at

  1. Primary structure of a key enzyme in plant tetrapyrrole synthesis: glutamate 1-semialdehyde aminotransferase.

    PubMed Central

    Grimm, B

    1990-01-01

    The formation of delta-aminolevulinate from glutamate 1-semialdehyde (GSA) is catalyzed by glutamate 1-semialdehyde aminotransferase (EC 5.4.3.8). The active form of the barley enzyme appears to be a dimer of identical subunits with a molecular mass of 46 kDa. From the purified enzyme, amino acid sequences of the N-terminal ends of the mature protein as well as an internal peptide were determined. DNA primers deduced from these peptide sequences were used to amplify with the polymerase chain reaction a cDNA sequence encoding part of the enzyme. Screening a cDNA library with this DNA fragment identified a full-length clone encoding the 49,540-Da precursor of the GSA aminotransferase. The transit peptide for chloroplast import consists of 34 amino acids. GSA aminotransferase and a precursor form were expressed on a multicopy plasmid in Escherichia coli. Both recombinant gene products reacted with an antibody against the barley GSA aminotransferase. Active barley GSA aminotransferase expressed in E. coli was shown to be active in assays of bacterial cell extracts. As a gene symbol for barley GSA aminotransferase, Gsa is proposed. Images PMID:2349227

  2. Kinemage of action - Proposed reaction mechanism of glutamate-1-semialdehyde aminomutase at an atomic level

    SciTech Connect

    Sorensen, John L.; Stetefeld, Joerg

    2011-10-07

    Highlights: {yields} Inhibitors of tetrapyrrole cofactor biosynthesis may be useful antibiotics. {yields} Mechanism of critical enzyme, glutamate-1-semialdehyde aminomutase, is presented. {yields} Unique vitamin B6-dependant enzyme traps intermediate in active site. {yields} Molecular dynamics show that a re-orientation of the substrate is required. -- Abstract: Glutamate-1-semialdehyde aminomutase (GSAM), a key enzyme in tetrapyrrole cofactor biosynthesis, performs a unique transamination on a single substrate. The substrate, glutamate-1-semialdehyde (GSA), undergoes a reaction that exchanges the position of an amine and a carbonyl group to produce 5-aminolevulinic acid (ALA). This transamination reaction is unique in the fact that is does not require an external cofactor to act as a nitrogen donor or acceptor in this transamination reaction. One of the other remarkable features of the catalytic mechanism is the release free in the enzyme active site of the intermediate 4,5-diaminovaleric acid (DAVA). The action of a gating loop prevents the escape of DAVA from the active site. In a MD simulation approach, using snapshots provided by X-ray crystallography and protein crystal absorption spectrometry data, the individual catalytic steps in this unique intramolecular transamination have been elucidated.

  3. Crystal structure of glutamate-1-semialdehyde-2,1-aminomutase from Arabidopsis thaliana

    SciTech Connect

    Song, Yingxian; Pu, Hua; Jiang, Tian; Zhang, Lixin; Ouyang, Min

    2016-05-23

    A structural study of A. thaliana glutamate-1-semialdehyde-2,1-aminomutase (GSAM) has revealed asymmetry in cofactor binding as well as in the gating-loop orientation, which supports the previously proposed negative cooperativity between monomers of GSAM. Glutamate-1-semialdehyde-2,1-aminomutase (GSAM) catalyzes the isomerization of glutamate-1-semialdehyde (GSA) to 5-aminolevulinate (ALA) and is distributed in archaea, most bacteria and plants. Although structures of GSAM from archaea and bacteria have been resolved, a GSAM structure from a higher plant is not available, preventing further structure–function analysis. Here, the structure of GSAM from Arabidopsis thaliana (AtGSA1) obtained by X-ray crystallography is reported at 1.25 Å resolution. AtGSA1 forms an asymmetric dimer and displays asymmetry in cofactor binding as well as in the gating-loop orientation, which is consistent with previously reported Synechococcus GSAM structures. While one monomer binds PMP with the gating loop fixed in the open state, the other monomer binds either PMP or PLP and the gating loop is ready to close. The data also reveal the mobility of residues Gly163, Ser164 and Gly165, which are important for reorientation of the gating loop. Furthermore, the asymmetry of the AtGSA1 structure supports the previously proposed negative cooperativity between monomers of GSAM.

  4. AGXT2: a promiscuous aminotransferase

    PubMed Central

    Rodionov, Roman N.; Jarzebska, Natalia; Weiss, Norbert; Lentz, Steven R.

    2014-01-01

    Alanine-glyoxylate aminotransferase 2 (AGXT2) is a multifunctional mitochondrial aminotransferase that was first identified in 1978. The physiological importance of AGXT2 was largely overlooked for three decades because AGXT2 is less active in glyoxylate metabolism than AGXT1, the enzyme that is deficient in primary hyperoxaluria type I. Recently, several novel functions of AGXT2 have been “rediscovered” in the setting of modern genomic and metabolomic studies. It is now apparent that AGXT2 has multiple substrates and products and that altered AGXT2 activity may contribute to the pathogenesis of cardiovascular, renal, neurological and hematological diseases. This article reviews the biochemical properties and physiological functions of AGXT2, its unique role at the intersection of key mitochondrial pathways, and its potential as a drug target. PMID:25294000

  5. Intramitochondrial localization of alanine aminotransferase in rat-liver mitochondria: comparison with glutaminase and aspartate aminotransferase.

    PubMed

    Masola, B; Devlin, T M

    1995-12-01

    The removal of the outer mitochondrial membrane and hence of constituents of the intermembrane space in rat-liver mitochondria using digitonin showed that phosphate-dependent glutaminase, alanine and aspartate aminotransferase were localized in the mitoplasts. Further fractionation of mitoplasts following their sonication resulted in 90% of glutaminase, 98% of alanine aminotransferase and 48% of aspartate aminotransferase being recovered in the soluble fraction while the remainder of each enzyme was recovered in the sonicated vesicles fraction. These results indicated that glutaminase and alanine aminotransferase were soluble matrix enzymes, the little of each enzyme recovered in the sonicated vesicles fraction being probably due to entrapment in the vesicles. Aspartate aminotransferase had dual localization, in the inner membrane and matrix with the high specific activity in sonicated vesicles confirming its association with the membrane. Activation experiments suggested that the membrane-bound enzyme was localized on the inner side of the inner mitochondrial membrane.

  6. Plasma aminotransferase concentrations in preterm infants.

    PubMed

    Victor, S; Dickinson, H; Turner, M A

    2011-03-01

    The aim of this study was to generate reference ranges for aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in preterm infants by describing the observed plasma concentration of these enzymes in babies born between 22 and 36 weeks' gestation. A service evaluation was conducted in babies admitted to two large neonatal intensive care units in the UK. 7006 blood samples from 1860 infants admitted to the two units between 2004 and 2008 were included. Extremely premature infants had high plasma enzyme activities when compared to babies at a later corrected gestational age. This may be due to more severe illness immediately after birth.

  7. Alanine aminotransferase controls seed dormancy in barley

    PubMed Central

    Sato, Kazuhiro; Yamane, Miki; Yamaji, Nami; Kanamori, Hiroyuki; Tagiri, Akemi; Schwerdt, Julian G.; Fincher, Geoffrey B.; Matsumoto, Takashi; Takeda, Kazuyoshi; Komatsuda, Takao

    2016-01-01

    Dormancy allows wild barley grains to survive dry summers in the Near East. After domestication, barley was selected for shorter dormancy periods. Here we isolate the major seed dormancy gene qsd1 from wild barley, which encodes an alanine aminotransferase (AlaAT). The seed dormancy gene is expressed specifically in the embryo. The AlaAT isoenzymes encoded by the long and short dormancy alleles differ in a single amino acid residue. The reduced dormancy allele Qsd1 evolved from barleys that were first domesticated in the southern Levant and had the long dormancy qsd1 allele that can be traced back to wild barleys. The reduced dormancy mutation likely contributed to the enhanced performance of barley in industrial applications such as beer and whisky production, which involve controlled germination. In contrast, the long dormancy allele might be used to control pre-harvest sprouting in higher rainfall areas to enhance global adaptation of barley. PMID:27188711

  8. [Conformation of aspartate aminotransferase in crystals].

    PubMed

    Borisov, V V; Borisova, S N; Sosfenov, N I; Dikson, Kh BF

    1983-01-01

    X-ray study of chicken cytosolic aspartate aminotransferase revealed conformational changes in the protein of two kinds: (1) a shift of the small domain adjacent to substrate-binding area due to interaction of the protein with two carboxyl groups of substrate and (2) a change in inclination of the coenzyme plane due to replacement of C = N bond of the coenzyme with Lys-258 by C = N bond with a substrate. An asymmetry in subunit behaviour is observed in both cases: the domain is shifted in one subunit and the coenzyme is rotated in other. Substrate-binding properties of each subunit are strictly dependent on the protein conformation in substrate-binding area.

  9. Crystal Structure of Human Kynurenine Aminotransferase ll*

    SciTech Connect

    Han,Q.; Robinson, H.; Li, J.

    2008-01-01

    Human kynurenine aminotransferase II (hKAT-II) efficiently catalyzes the transamination of knunrenine to kynurenic acid (KYNA). KYNA is the only known endogenous antagonist of N-methyl-d-aspartate (NMDA) receptors and is also an antagonist of 7-nicotinic acetylcholine receptors. Abnormal concentrations of brain KYNA have been implicated in the pathogenesis and development of several neurological and psychiatric diseases in humans. Consequently, enzymes involved in the production of brain KYNA have been considered potential regulatory targets. In this article, we report a 2.16 Angstroms crystal structure of hKAT-II and a 1.95 Angstroms structure of its complex with kynurenine. The protein architecture of hKAT-II reveals that it belongs to the fold-type I pyridoxal 5-phosphate (PLP)-dependent enzymes. In comparison with all subclasses of fold-type I-PLP-dependent enzymes, we propose that hKAT-II represents a novel subclass in the fold-type I enzymes because of the unique folding of its first 65 N-terminal residues. This study provides a molecular basis for future effort in maintaining physiological concentrations of KYNA through molecular and biochemical regulation of hKAT-II.

  10. Conversion of cysteine to 3-mercaptopyruvic acid by bacterial aminotransferases.

    PubMed

    Andreeßen, Christina; Gerlt, Vanessa; Steinbüchel, Alexander

    2017-04-01

    3-Mercaptopyruvate (3MPy), a structural analog of 3-mercaptopropionic acid, is a precursor compound for biosynthesis of polythioesters in bacteria. The cost-effectiveness and sustainability of the whole process could be greatly improved by using the cysteine degradation pathway for an intracellular supply of 3MPy. Transamination of cysteine to its corresponding α-keto acid 3MPy is catalyzed by cysteine aminotransferases (CAT). However, CAT activity has so far not been described for bacterial aminotransferases (AT), and it was unknown whether they can be applied for the conversion of cysteine to 3MPy. In this study, we selected eight bacterial aminotransferases based on sequence homology to CAT of Rattus norvegicus (Got1). The aminotransferases included four aspartate aminotransferases (AATs) and four aromatic amino acid aminotransferases (ArATs) from Advenella mimigardefordensis DPN7, Escherichia coli MG1655, Shimwellia blattae ATCC 33430, Ralstonia eutropha H16 and Paracoccus denitrificans PD1222. For a more detailed characterization, all selected AAT or ArAT encoding genes were heterologously expressed in E. coli and purified. CAT activity was detected for all aminotransferases when a novel continuous coupled enzyme assay was applied. Kinetic studies revealed the highest catalytic efficiency of 5.1mM/s for AAT from A. mimigardefordensis. Formation of 3MPy from cysteine could additionally be verified by an optimized approach using derivatization of 3MPy with the Girard T reagent and liquid chromatography-mass spectrometry analyses.

  11. Similarities between cysteinesulphinate transaminase and aspartate aminotransferase.

    PubMed

    Recasens, M; Mandel, P

    1979-01-01

    A method for the purification of two cysteinesulphinate transaminases, A and B (EC 2.6.1), is described. These enzymes catalyse the conversion of cysteinesulphinic acid to beta-sulphinyl pyruvate. The final preparations are homogeneous by polyacrylamide gel electrophoresis, sodium dodecyl sulphate-polyacrylamide gel electrophoresis and isoelectrofocusing. The molecular weight of the subunits is 41 000 for cysteinesulphinate transaminase A and 43 400 for B. Both enzymes are unspecific, as L-asparate, L-glutamate and L-cysteic acid serve as substrates in addition to L-cysteinesulphinic acid. Cysteinesulphinate transaminase A has a Km of 9.8 mM for cysteinesulphinic acid and 0.25 mM for aspartic acid, whereas the B enzyme has a Km of 6.5 mM for cysteinesulphinic acid and 1.4 mM for aspartic acid. The Vmax values of the A and B enzymes are respectively 7.1 and 6.2 mmol h-1 mg-1 protein for aspartic acid and 45 and 9.3 mmol h-1 mg-1 protein for cysteinesulphinic acid. Both enzymes exhibit maximum activity at pH 8.6. A high specific activity is found in optimal conditions for these two transaminases, the pI values being 9.06 and 5.70 for cysteinesulphinate transaminase A and B respectively. These results have been compared with those already obtained for purified aspartate aminotransferase. Similarities in the pathways of taurine and gamma-aminobutyric acid (GABA) metabolism are discussed.

  12. Kynurenine Aminotransferase Isozyme Inhibitors: A Review

    PubMed Central

    Nematollahi, Alireza; Sun, Guanchen; Jayawickrama, Gayan S.; Church, W. Bret

    2016-01-01

    Kynurenine aminotransferase isozymes (KATs 1–4) are members of the pyridoxal-5’-phosphate (PLP)-dependent enzyme family, which catalyse the permanent conversion of l-kynurenine (l-KYN) to kynurenic acid (KYNA), a known neuroactive agent. As KATs are found in the mammalian brain and have key roles in the kynurenine pathway, involved in different categories of central nervous system (CNS) diseases, the KATs are prominent targets in the quest to treat neurodegenerative and cognitive impairment disorders. Recent studies suggest that inhibiting these enzymes would produce effects beneficial to patients with these conditions, as abnormally high levels of KYNA are observed. KAT-1 and KAT-3 share the highest sequence similarity of the isozymes in this family, and their active site pockets are also similar. Importantly, KAT-2 has the major role of kynurenic acid production (70%) in the human brain, and it is considered therefore that suitable inhibition of this isozyme would be most effective in managing major aspects of CNS diseases. Human KAT-2 inhibitors have been developed, but the most potent of them, chosen for further investigations, did not proceed in clinical studies due to the cross toxicity caused by their irreversible interaction with PLP, the required cofactor of the KAT isozymes, and any other PLP-dependent enzymes. As a consequence of the possibility of extensive undesirable adverse effects, it is also important to pursue KAT inhibitors that reversibly inhibit KATs and to include a strategy that seeks compounds likely to achieve substantial interaction with regions of the active site other than the PLP. The main purpose of this treatise is to review the recent developments with the inhibitors of KAT isozymes. This treatise also includes analyses of their crystallographic structures in complex with this enzyme family, which provides further insight for researchers in this and related studies. PMID:27314340

  13. Substrate specificity and structure of human aminoadipate aminotransferase/kynurenine aminotransferase II.

    PubMed

    Han, Qian; Cai, Tao; Tagle, Danilo A; Robinson, Howard; Li, Jianyong

    2008-08-01

    KAT (kynurenine aminotransferase) II is a primary enzyme in the brain for catalysing the transamination of kynurenine to KYNA (kynurenic acid). KYNA is the only known endogenous antagonist of the N-methyl-D-aspartate receptor. The enzyme also catalyses the transamination of aminoadipate to alpha-oxoadipate; therefore it was initially named AADAT (aminoadipate aminotransferase). As an endotoxin, aminoadipate influences various elements of glutamatergic neurotransmission and kills primary astrocytes in the brain. A number of studies dealing with the biochemical and functional characteristics of this enzyme exist in the literature, but a systematic assessment of KAT II addressing its substrate profile and kinetic properties has not been performed. The present study examines the biochemical and structural characterization of a human KAT II/AADAT. Substrate screening of human KAT II revealed that the enzyme has a very broad substrate specificity, is capable of catalysing the transamination of 16 out of 24 tested amino acids and could utilize all 16 tested alpha-oxo acids as amino-group acceptors. Kinetic analysis of human KAT II demonstrated its catalytic efficiency for individual amino-group donors and acceptors, providing information as to its preferred substrate affinity. Structural analysis of the human KAT II complex with alpha-oxoglutaric acid revealed a conformational change of an N-terminal fraction, residues 15-33, that is able to adapt to different substrate sizes, which provides a structural basis for its broad substrate specificity.

  14. Biochemical and structural characterization of mouse mitochondrial aspartate aminotransferase, a newly identified kynurenine aminotransferase-IV

    PubMed Central

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A.; Li, Jianyong

    2010-01-01

    Synopsis Mammalian mitochondrial aspartate aminotransferase (mAspAT) is recently reported to have kynurenine aminotransferase (KAT) activity and plays a role in the biosynthesis of kynurenic acid (KYNA) in rat, mouse and human brains. This study concerns the biochemical and structural characterization of mouse mAspAT. In this study, mouse mAspAT cDNA was amplified from mouse brain first stand cDNA and its recombinant protein was expressed in an Escherichia coli expression system. Sixteen keto acids were tested for the co-substrate specificity of mouse mAspAT and fourteen of them were shown to be capable of serving as co-substrates for the enzyme. Structural analysis of mAspAT by macromolecular crystallography revealed that the cofactor binding residues of mAspAT are similar to those of other KATs. The substrate binding residues of mAspAT are slightly different from those of other KATs. Our data provide a biochemical and structural basis towards understanding the overall physiological role of mAspAT in vivo and insight into controlling the levels of endogenous KYNA through modulation of the enzyme in the mouse brain. PMID:20977429

  15. Substrate specificity and structure of human aminoadipate aminotransferase/kynurenine aminotransferase II

    PubMed Central

    HAN, Qian; CAI, Tao; TAGLE, Danilo A.; ROBINSON, Howard; LI, Jianyong

    2008-01-01

    Synopsis KAT (kynurenine aminotransferase) II is a primary enzyme in the brain for catalysing the transamination of kynurenine to KYNA (kynurenic acid). KYNA is the only known endogenous antagonist of the N-methyl-d-aspartate receptor. The enzyme also catalyses the transamination of aminoadipate to α-oxoadipate; therefore it was initially named AADAT (aminoadipate aminotransferase). As an endotoxin, aminoadipate influences various elements of glutamatergic neurotransmission and kills primary astrocytes in the brain. A number of studies dealing with the biochemical and functional characteristics of this enzyme exist in the literature, but a systematic assessment of KAT II addressing its substrate profile and kinetic properties has not been performed. The present study examines the biochemical and structural characterization of a human KAT II/AADAT. Substrate screening of human KAT II revealed that the enzyme has a very broad substrate specificity, is capable of catalysing the transamination of 16 out of 24 tested amino acids and could utilize all 16 tested α-oxo acids as amino-group acceptors. Kinetic analysis of human KAT II demonstrated its catalytic efficiency for individual amino-group donors and acceptors, providing information as to its preferred substrate affinity. Structural analysis of the human KAT II complex with α-oxoglutaric acid revealed a conformational change of an N-terminal fraction, residues 15-33, that is able to adapt to different substrate sizes, which provides a structural basis for its broad substrate specificity. PMID:18620547

  16. Biochemical and structural characterization of mouse mitochondrial aspartate aminotransferase, a newly identified kynurenine aminotransferase-IV.

    PubMed

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A; Li, Jianyong

    2011-10-01

    Mammalian mAspAT (mitochondrial aspartate aminotransferase) is recently reported to have KAT (kynurenine aminotransferase) activity and plays a role in the biosynthesis of KYNA (kynurenic acid) in rat, mouse and human brains. This study concerns the biochemical and structural characterization of mouse mAspAT. In this study, mouse mAspAT cDNA was amplified from mouse brain first stand cDNA and its recombinant protein was expressed in an Escherichia coli expression system. Sixteen oxo acids were tested for the co-substrate specificity of mouse mAspAT and 14 of them were shown to be capable of serving as co-substrates for the enzyme. Structural analysis of mAspAT by macromolecular crystallography revealed that the cofactor-binding residues of mAspAT are similar to those of other KATs. The substrate-binding residues of mAspAT are slightly different from those of other KATs. Our results provide a biochemical and structural basis towards understanding the overall physiological role of mAspAT in vivo and insight into controlling the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  17. Biochemical and structural characterization of mouse mitochondrial aspartate aminotransferase, a newly identified kynurenine aminotransferase-IV

    SciTech Connect

    Han, Q.; Robinson, H.; Cai, T.; Tagle, D. A.; Li, J.

    2011-10-01

    Mammalian mAspAT (mitochondrial aspartate aminotransferase) is recently reported to have KAT (kynurenine aminotransferase) activity and plays a role in the biosynthesis of KYNA (kynurenic acid) in rat, mouse and human brains. This study concerns the biochemical and structural characterization of mouse mAspAT. In this study, mouse mAspAT cDNA was amplified from mouse brain first stand cDNA and its recombinant protein was expressed in an Escherichia coli expression system. Sixteen oxo acids were tested for the co-substrate specificity of mouse mAspAT and 14 of them were shown to be capable of serving as co-substrates for the enzyme. Structural analysis of mAspAT by macromolecular crystallography revealed that the cofactor-binding residues of mAspAT are similar to those of other KATs. The substrate-binding residues of mAspAT are slightly different from those of other KATs. Our results provide a biochemical and structural basis towards understanding the overall physiological role of mAspAT in vivo and insight into controlling the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  18. Substrate Specificity and Structure of Human aminoadipate aminotransferase/kynurenine aminotransferase II

    SciTech Connect

    Han, Q.; Cai, T; Tagle, D; Robinson, H; Li, J

    2009-01-01

    KAT (kynurenine aminotransferase) II is a primary enzyme in the brain for catalysing the transamination of kynurenine to KYNA (kynurenic acid). KYNA is the only known endogenous antagonist of the N-methyl-D-aspartate receptor. The enzyme also catalyses the transamination of aminoadipate to alpha-oxoadipate; therefore it was initially named AADAT (aminoadipate aminotransferase). As an endotoxin, aminoadipate influences various elements of glutamatergic neurotransmission and kills primary astrocytes in the brain. A number of studies dealing with the biochemical and functional characteristics of this enzyme exist in the literature, but a systematic assessment of KAT II addressing its substrate profile and kinetic properties has not been performed. The present study examines the biochemical and structural characterization of a human KAT II/AADAT. Substrate screening of human KAT II revealed that the enzyme has a very broad substrate specificity, is capable of catalysing the transamination of 16 out of 24 tested amino acids and could utilize all 16 tested alpha-oxo acids as amino-group acceptors. Kinetic analysis of human KAT II demonstrated its catalytic efficiency for individual amino-group donors and acceptors, providing information as to its preferred substrate affinity. Structural analysis of the human KAT II complex with alpha-oxoglutaric acid revealed a conformational change of an N-terminal fraction, residues 15-33, that is able to adapt to different substrate sizes, which provides a structural basis for its broad substrate specificity.

  19. Substrate Specificity and Structure of Human Aminoadipate Aminotransferase/kynurenine Aminotransferase II

    SciTech Connect

    Han,Q.; Cai, T.; Tagle, D.; Robinson, H.; Li, J.

    2008-01-01

    KAT (kynurenine aminotransferase) II is a primary enzyme in the brain for catalysing the transamination of kynurenine to KYNA (kynurenic acid). KYNA is the only known endogenous antagonist of the N-methyl-D-aspartate receptor. The enzyme also catalyses the transamination of aminoadipate to a-oxoadipate; therefore it was initially named AADAT (aminoadipate aminotransferase). As an endotoxin, aminoadipate influences various elements of glutamatergic neurotransmission and kills primary astrocytes in the brain. A number of studies dealing with the biochemical and functional characteristics of this enzyme exist in the literature, but a systematic assessment of KAT II addressing its substrate profile and kinetic properties has not been performed. The present study examines the biochemical and structural characterization of a human KAT II/AADAT. Substrate screening of human KAT II revealed that the enzyme has a very broad substrate specificity, is capable of catalysing the transamination of 16 out of 24 tested amino acids and could utilize all 16 tested a-oxo acids as amino-group acceptors. Kinetic analysis of human KAT II demonstrated its catalytic efficiency for individual amino-group donors and acceptors, providing information as to its preferred substrate affinity. Structural analysis of the human KAT II complex with a-oxoglutaric acid revealed a conformational change of an N-terminal fraction, residues 15-33, that is able to adapt to different substrate sizes, which provides a structural basis for its broad substrate specificity.

  20. Observations of alanine aminotransferase and aspartate aminotransferase in THRIVE studies treated orally with ximelagatran.

    PubMed

    Harenberg, Job; Jörg, Ingrid; Weiss, Christel

    2006-01-01

    Treatment of acute venous thromboembolism (VTE) and prophylaxis of recurrent events has been investigated in the THRIVE (THRombin Inhibitor in Venous Thrombe Embolism) Treatment and the THRIVE III trial using the oral direct thrombin inhibitor ximelagatran. Alanine aminotransferase (ALAT) increased in 9.6% and 6.4% of patients in the THRIVE Treatment and THRIVE III trials, respectively. The authors analysed the time course of the ALAT and in additionally of aspartate aminotransferase (ASAT) in blood from 52 and 23 patients participating in the THRIVE Treatment and the THRIVE III trials in Germany. Analysis of variance for repeated measures and t test were performed. In the THRIVE Treatment trial, ALAT was significantly higher at week 2 for enoxaparin/warfarin (p => .0039, t test) and at months 3 and 6 for ximelagatran (p = .0453, p = .0014, respectively). ASAT and ASAT/ALAT ratio values did not increase and not differ for both groups. In the THRIVE III trial, ALAT and ASAT did not increase and did not differ compared to the comparator placebo. 2 x 36 mg Ximelagatran, induced higher ALAT values at months 3 and 6 compared to 2 x 24 mg ximelagatran (p = .0105, p = .0063, respectively). ASAT did not differ between the two doses of ximelagatran. The ASAT/ALAT ratios were lower at week 2 for enoxaparin/warfarin (t-test, p = .0032) and at month 3 and 6 for 2 x 36 mg versus warfarin or 2 x 24 mg Ximelagatran (p between .0187 and .0002). The authors conclude that ALAT increases dose dependently during therapy with ximelagatran. The less frequent and lower increase of ASAT values compared to ALAT values indicates a nontoxic effect of ximelagatran on liver cells.

  1. Correlation of serum alanine aminotransferase and aspartate aminotransferase with coronary heart disease

    PubMed Central

    Shen, Jianying; Zhang, Jingying; Wen, Jing; Ming, Qiang; Zhang, Ji; Xu, Yawei

    2015-01-01

    Objective: This study aimed to explore the relationship between different risk factors (especially serum alanine aminotransferase [ALT] and aspartate aminotransferase [AST]) and coronary heart disease (CHD). Methods: A total of 610 inpatients were recruited. Initial coronary angiography (CAG) was performed to evaluate the severity of coronary lesions. On the basis of findings from CAG, patients were divided into control group (n=260) and CHD group (n=350). Logistic regression analysis was employed for the evaluation of clinical characteristics and biochemical parameters, aiming to explore the relationship between risk factors (including AST and ALT) and CHD. Results: Results showed type 2 diabetes, hypertension, dyslipidemia, smoking and family history of CHD were clinical risk factors of CHD. Laboratory examinations showed the serum levels of triglycerides, low-density lipoprotein, AST and ALT in CHD group were significantly higher than those in control group (P<0.05). Of these parameters, the AST was 50.98±8.12 U/L in CHD group and 20.14±3.94 U/L in control group (P<0.01); the ALT was 42.31±8.34 U/L in CHD group and 18.25±6.38 U/L in control group (P<0.01). Conclusion: The serum levels of AST and ALT in CHD patients are higher than those in controls. High serum AST and ALT are biochemical markers which can be used to predict the severity of CHD and are also independent risk factors of CHD. PMID:26064360

  2. Molecular cloning, expression and characterization of pyridoxamine–pyruvate aminotransferase

    PubMed Central

    Yoshikane, Yu; Yokochi, Nana; Ohnishi, Kouhei; Hayashi, Hideyuki; Yagi, Toshiharu

    2006-01-01

    Pyridoxamine–pyruvate aminotransferase is a PLP (pyridoxal 5′-phosphate) (a coenzyme form of vitamin B6)-independent aminotransferase which catalyses a reversible transamination reaction between pyridoxamine and pyruvate to form pyridoxal and L-alanine. The gene encoding the enzyme has been identified, cloned and overexpressed for the first time. The mlr6806 gene on the chromosome of a symbiotic nitrogen-fixing bacterium, Mesorhizobium loti, encoded the enzyme, which consists of 393 amino acid residues. The primary sequence was identical with those of archaeal aspartate aminotransferase and rat serine–pyruvate aminotransferase, which are PLP-dependent aminotransferases. The results of fold-type analysis and the consensus amino acid residues found around the active-site lysine residue identified in the present study showed that the enzyme could be classified into class V aminotransferases of fold type I or the AT IV subfamily of the α family of the PLP-dependent enzymes. Analyses of the absorption and CD spectra of the wild-type and point-mutated enzymes showed that Lys197 was essential for the enzyme activity, and was the active-site lysine residue that corresponded to that found in the PLP-dependent aminotransferases, as had been suggested previously [Hodsdon, Kolb, Snell and Cole (1978) Biochem. J. 169, 429–432]. The Kd value for pyridoxal determined by means of CD was 100-fold lower than the Km value for it, suggesting that Schiff base formation between pyridoxal and the active-site lysine residue is partially rate determining in the catalysis of pyridoxal. The active-site structure and evolutionary aspects of the enzyme are discussed. PMID:16545075

  3. Structure of putrescine aminotransferase from Escherichia coli provides insights into the substrate specificity among class III aminotransferases.

    PubMed

    Cha, Hyung Jin; Jeong, Jae-Hee; Rojviriya, Catleya; Kim, Yeon-Gil

    2014-01-01

    YgjG is a putrescine aminotransferase enzyme that transfers amino groups from compounds with terminal primary amines to compounds with an aldehyde group using pyridoxal-5'-phosphate (PLP) as a cofactor. Previous biochemical data show that the enzyme prefers primary diamines, such as putrescine, over ornithine as a substrate. To better understand the enzyme's substrate specificity, crystal structures of YgjG from Escherichia coli were determined at 2.3 and 2.1 Å resolutions for the free and putrescine-bound enzymes, respectively. Sequence and structural analyses revealed that YgjG forms a dimer that adopts a class III PLP-dependent aminotransferase fold. A structural comparison between YgjG and other class III aminotransferases revealed that their structures are similar. However, YgjG has an additional N-terminal helical structure that partially contributes to a dimeric interaction with the other subunit via a helix-helix interaction. Interestingly, the YgjG substrate-binding site entrance size and charge distribution are smaller and more hydrophobic than other class III aminotransferases, which suggest that YgjG has a unique substrate binding site that could accommodate primary aliphatic diamine substrates, including putrescine. The YgjG crystal structures provide structural clues to putrescine aminotransferase substrate specificity and binding.

  4. METHANOGENS WITH PSEUDOMUREIN USE DIAMINOPIMELATE AMINOTRANSFERASE IN LYSINE BIOSYNTHESIS

    PubMed Central

    Graham, David E.; Huse, Holly K.

    2008-01-01

    Methanothermobacter thermautotrophicus uses lysine for both protein synthesis and cross-linking pseudomurein in its cell wall. A diaminopimelate aminotransferase enzyme from this methanogen (MTH0052) converts tetrahydrodipicolinate to L,L-diaminopimelate, a lysine precursor. This gene complemented an Escherichia coli diaminopimelate auxotrophy, and the purified protein catalyzed the transamination of diaminopimelate to tetrahydrodipicolinate. Phylogenetic analysis indicated this gene was recruited from anaerobic Gram-positive bacteria. These results expand the family of diaminopimelate aminotransferases to a diverse set of plant, bacterial and archaeal homologs. In contrast marine methanogens from the Methanococcales, which lack pseudomurein, appear to use a different diaminopimelate pathway for lysine biosynthesis. PMID:18371309

  5. Biocatalytic potential of vanillin aminotransferase from Capsicum chinense

    PubMed Central

    2014-01-01

    Background The conversion of vanillin to vanillylamine is a key step in the biosynthetic route towards capsaicinoids in pungent cultivars of Capsicum sp. The reaction has previously been annotated to be catalysed by PAMT (putative aminotransferase; [GenBank: AAC78480.1, Swiss-Prot: O82521]), however, the enzyme has previously not been biochemically characterised in vitro. Results The biochemical activity of the transaminase was confirmed by direct measurement of the reaction with purified recombinant enzyme. The enzyme accepted pyruvate, and oxaloacetate but not 2-oxoglutarate as co-substrate, which is in accordance with other characterised transaminases from the plant kingdom. The enzyme was also able to convert (S)-1-phenylethylamine into acetophenone with high stereo-selectivity. Additionally, it was shown to be active at a broad pH range. Conclusions We suggest PAMT to be renamed to VAMT (vanillin aminotransferase, abbreviation used in this study) as formation of vanillin from vanillylamine could be demonstrated. Furthermore, due to high stereoselectivity and activity at physiological pH, VAMT is a suitable candidate for biocatalytic transamination in a recombinant whole-cell system. PMID:24712445

  6. Serum γ-Glutamyltransferase, Alanine Aminotransferase and Aspartate Aminotransferase Activity in Healthy Blood Donor of Different Ethnic Groups in Gorgan

    PubMed Central

    Mehrpouya, Masoumeh; Pourhashem, Zeinab

    2016-01-01

    Introduction Measure of liver enzymes may help to increase safety of blood donation for both blood donor and recipient. Determination of liver enzymes may prepare valuable clinical information. Aim To assess serum γ-Glutamyltransferase (GGT), Alanine Aminotransferase (ALT), and Aspartate Aminotransferase (AST) activities in healthy blood donors in different ethnic groups in Gorgan. Materials and Methods This study was performed in 450 healthy male blood donors, in three ethnic groups (Fars, Sistanee and Turkman) who attended Gorgan blood transfusion center. Liver enzymes (GGT, ALT and AST) were determined. Results Serum AST and ALT in three ethnic groups were significant except for serum GGT levels. There was significant correlation between family histories of liver disease and systolic blood pressure and AST in Fars, and GGT in Sistanee ethnic groups. Conclusion Several factors, such as age, family history of diabetes mellitus, family history of liver disease and smoking habit had no effect on some liver enzymes in different ethnic groups in this area. Variation of AST, ALT, and GGT enzyme activities in healthy subjects was associated with some subjects in our study groups. According to our study, it suggests that screening of AST and GGT enzymes in subjects with family history of liver disease is necessary in different ethnic groups. PMID:27630834

  7. Synthesis and evaluation of novel heteroaromatic substrates of GABA aminotransferase

    PubMed Central

    Hawker, Dustin D.; Silverman, Richard B.

    2012-01-01

    Two principal neurotransmitters are involved in the regulation of mammalian neuronal activity, namely, γ-aminobutyric acid (GABA), an inhibitory neurotransmitter, and L-glutamic acid, an excitatory neurotransmitter. Low GABA levels in the brain have been implicated in epilepsy and several other neurological diseases. Because of GABA’s poor ability to cross the blood-brain barrier (BBB), a successful strategy to raise brain GABA concentrations is the use of a compound that does cross the BBB and inhibits or inactivates GABA aminotransferase (GABA-AT), the enzyme responsible for GABA catabolism. Vigabatrin, a mechanism-based inactivator of GABA-AT, is currently a successful therapeutic for epilepsy, but has harmful side effects, leaving a need for improved GABA-AT inactivators. Here, we report the synthesis and evaluation of a series of heteroaromatic GABA analogues as substrates of GABA-AT, which will be used as the basis for the design of novel enzyme inactivators. PMID:22944334

  8. Ambient Ionization Mass Spectrometry Measurement of Aminotransferase Activity

    NASA Astrophysics Data System (ADS)

    Yan, Xin; Li, Xin; Zhang, Chengsen; Xu, Yang; Cooks, R. Graham

    2017-01-01

    A change in enzyme activity has been used as a clinical biomarker for diagnosis and is useful in evaluating patient prognosis. Current laboratory measurements of enzyme activity involve multi-step derivatization of the reaction products followed by quantitative analysis of these derivatives. This study simplified the reaction systems by using only the target enzymatic reaction and directly detecting its product. A protocol using paper spray mass spectrometry for identifying and quantifying the reaction product has been developed. Evaluation of the activity of aspartate aminotransferase (AST) was chosen as a proof-of-principle. The volume of sample needed is greatly reduced compared with the traditional method. Paper spray has a desalting effect that avoids sprayer clogging problems seen when examining serum samples by nanoESI. This very simple method does not require sample pretreatment and additional derivatization reactions, yet it gives high quality kinetic data, excellent limits of detection (60 ppb from serum), and coefficients of variation <10% in quantitation.

  9. Crystal Structures of Aedes Aegypt Alanine Glyoxylate Aminotransferase

    SciTech Connect

    Han,Q.; Robinson, H.; Gao, Y.; Vogelaar, N.; Wilson, S.; Rizzi, M.; Li, J.

    2006-01-01

    Mosquitoes are unique in having evolved two alanine glyoxylate aminotransferases (AGTs). One is 3-hydroxykynurenine transaminase (HKT), which is primarily responsible for catalyzing the transamination of 3-hydroxykynurenine (3-HK) to xanthurenic acid (XA). Interestingly, XA is used by malaria parasites as a chemical trigger for their development within the mosquito. This 3-HK to XA conversion is considered the major mechanism mosquitoes use to detoxify the chemically reactive and potentially toxic 3-HK. The other AGT is a typical dipteran insect AGT and is specific for converting glyoxylic acid to glycine. Here we report the 1.75{angstrom} high-resolution three-dimensional crystal structure of AGT from the mosquito Aedes aegypti (AeAGT) and structures of its complexes with reactants glyoxylic acid and alanine at 1.75 and 2.1{angstrom} resolution, respectively. This is the first time that the three-dimensional crystal structures of an AGT with its amino acceptor, glyoxylic acid, and amino donor, alanine, have been determined. The protein is dimeric and adopts the type I-fold of pyridoxal 5-phosphate (PLP)-dependent aminotransferases. The PLP co-factor is covalently bound to the active site in the crystal structure, and its binding site is similar to those of other AGTs. The comparison of the AeAGT-glyoxylic acid structure with other AGT structures revealed that these glyoxylic acid binding residues are conserved in most AGTs. Comparison of the AeAGT-alanine structure with that of the Anopheles HKT-inhibitor complex suggests that a Ser-Asn-Phe motif in the latter may be responsible for the substrate specificity of HKT enzymes for 3-HK.

  10. Salivary lactate dehydrogenase and aminotransferases in diabetic patients

    PubMed Central

    Malicka, Barbara; Skoskiewicz-Malinowska, Katarzyna; Kaczmarek, Urszula

    2016-01-01

    Abstract Diabetes mellitus (DM) is a group of metabolic diseases resulting from impaired insulin secretion and/or action. DM is characterized by hyperglycemia that can lead to the dysfunction or damage of organs, including the salivary glands. The aim of this study was to compare the levels of salivary lactate dehydrogenase (LDH), aspartate aminotransferase (AST), and alanine aminotransferase (ALT) in diabetic patients. The study was approved by the Bioethics Committee of Wroclaw Medical University (Poland). The study comprised 90 adults of both sexes, aged 21 to 57 years. The patients were divided into 3 groups: type 1 diabetics (D1), type 2 diabetics (D2), and a healthy control group (C). Each group consisted of 30 age- and sex-matched subjects. Total protein (P, by Lowry method), LDH, AST, ALT (with Alpha Diagnostics kits), and salivary flow rate were measured in unstimulated mixed saliva. The level of glycosylated hemoglobin (HbA1c) was measured with DCA 2000 Reagent Kit. The obtained data were analyzed using the Mann–Whitney U test and the Spearman rank at a significance level of P < 0.05 with the use of STATISTICA 9.0 software. In comparison with C, D1 presented a significantly higher activity of LDH (P < 0.001), AST (P < 0.001), and ALT (P < 0.01), whereas D2 indicated higher levels of LDH (P < 0.001) and ALT (P < 0.05) compared with C. Comparing D1 to D2, approximately 3-fold higher activity of AST (P < 0.01) and approximately 4.5-fold higher activity of ALT (P < 0.01) was observed. Higher levels of salivary LDH, AST, and ALT in D1 compared with D2 and C confirm that salivary glands of D1 might be attributed to autoimmunological damage associated with the pathomechanism of DM. PMID:27893660

  11. Unique substrate specificity of ornithine aminotransferase from Toxoplasma gondii.

    PubMed

    Astegno, Alessandra; Maresi, Elena; Bertoldi, Mariarita; La Verde, Valentina; Paiardini, Alessandro; Dominici, Paola

    2017-03-07

    Toxoplasma gondii is a protozoan parasite of medical and veterinary relevance responsible for toxoplasmosis in humans. As an efficacious vaccine remains a challenge, chemotherapy is still the most effective way to combat the disease. In search of novel druggable targets, we performed a thorough characterization of the putative pyridoxal 5'-phosphate (PLP)-dependent enzyme ornithine aminotransferase from T. gondii ME49 (TgOAT). We overexpressed the protein in Escherichia coli and analysed its molecular and kinetic properties by UV-visible absorbance, fluorescence and CD spectroscopy, in addition to kinetic studies of both the steady state and pre-steady state. TgOAT is largely similar to OATs from other species regarding its general transamination mechanism and spectral properties of PLP; however, it does not show a specific ornithine aminotransferase activity like its human homologue, but exhibits both N-acetylornithine and γ-aminobutyric acid (GABA) transaminase activity in vitro, suggesting a role in both arginine and GABA metabolism in vivo The presence of Val79 in the active site of TgOAT in place of Tyr, as in its human counterpart, provides the necessary room to accommodate N-acetylornithine and GABA, resembling the active site arrangement of GABA transaminases. Moreover, mutation of Val79 to Tyr results in a change of substrate preference between GABA, N-acetylornithine and L-ornithine, suggesting a key role of Val79 in defining substrate specificity. The findings that TgOAT possesses parasite-specific structural features as well as differing substrate specificity from its human homologue make it an attractive target for anti-toxoplasmosis inhibitor design that can be exploited for chemotherapeutic intervention.

  12. Crystal structures of Aedes aegypti alanine glyoxylate aminotransferase.

    PubMed

    Han, Qian; Robinson, Howard; Gao, Yi Gui; Vogelaar, Nancy; Wilson, Scott R; Rizzi, Menico; Li, Jianyong

    2006-12-01

    Mosquitoes are unique in having evolved two alanine glyoxylate aminotransferases (AGTs). One is 3-hydroxykynurenine transaminase (HKT), which is primarily responsible for catalyzing the transamination of 3-hydroxykynurenine (3-HK) to xanthurenic acid (XA). Interestingly, XA is used by malaria parasites as a chemical trigger for their development within the mosquito. This 3-HK to XA conversion is considered the major mechanism mosquitoes use to detoxify the chemically reactive and potentially toxic 3-HK. The other AGT is a typical dipteran insect AGT and is specific for converting glyoxylic acid to glycine. Here we report the 1.75A high-resolution three-dimensional crystal structure of AGT from the mosquito Aedes aegypti (AeAGT) and structures of its complexes with reactants glyoxylic acid and alanine at 1.75 and 2.1A resolution, respectively. This is the first time that the three-dimensional crystal structures of an AGT with its amino acceptor, glyoxylic acid, and amino donor, alanine, have been determined. The protein is dimeric and adopts the type I-fold of pyridoxal 5-phosphate (PLP)-dependent aminotransferases. The PLP co-factor is covalently bound to the active site in the crystal structure, and its binding site is similar to those of other AGTs. The comparison of the AeAGT-glyoxylic acid structure with other AGT structures revealed that these glyoxylic acid binding residues are conserved in most AGTs. Comparison of the AeAGT-alanine structure with that of the Anopheles HKT-inhibitor complex suggests that a Ser-Asn-Phe motif in the latter may be responsible for the substrate specificity of HKT enzymes for 3-HK.

  13. The Relationships between Respiratory Virus Infection and Aminotransferase in Children

    PubMed Central

    Oh, Jun Suk; Choi, Jun Sik; Lee, Young Hyuk; Ko, Kyung Og; Lim, Jae Woo; Cheon, Eun Jung; Lee, Gyung Min

    2016-01-01

    Purpose We sought to examine the relationship between the clinical manifestations of nonspecific reactive hepatitis and respiratory virus infection in pediatric patients. Methods Patients admitted to the pediatric unit of Konyang University Hospital for lower respiratory tract disease between January 1, 2014 and December 31, 2014 and who underwent reverse transcriptase polymerase chain reaction tests were examined. The patients were divided into those with increased levels of alanine aminotransferase (ALT) or aspartate aminotransferase (AST) and those with normal ALT or AST levels. Further, patients with increased ALT and AST levels were individually compared with patients in the normal group, and the blood test results were compared according to the type of respiratory virus. Results Patients with increased ALT or AST levels had one more day of hospital stay, on average, compared with patients in the normal group (5.3±3.1 days vs. 4.4±3.0 days, p=0.019). Patients in the increased ALT level group were younger and had a longer mean hospital stay, compared with patients in the normal group (p=0.022 and 0.003, respectively). The incidences of increased ALT or AST were the highest in adenovirus infections (6/24, 25.0%), followed by enterovirus (2/11, 18.2%) and respiratory syncytial virus A (21/131, 16.0%) infections. Conclusion Nonspecific reactive hepatitis is more common among patients with adenovirus, enterovirus and respiratory syncytial virus infection, as well as among those infected at a younger age. Compared with AST levels, ALT levels are better indicators of the severity of nonspecific reactive hepatitis. PMID:28090469

  14. Characterisation of Potential Antimicrobial Targets in Bacillus spp. I. Aminotransferases and Methionine Regeneration in Bacillus subtilis

    DTIC Science & Technology

    2002-07-01

    targets in Bacillus spp. I. Aminotransferases and methionine regeneration in Bacillus subtilis. Bradley J. Berger and Marvin H. Knodel Defence R&D...Characterisation of potential antimicrobial targets in Bacillus spp. I. Aminotransferases and methionine regeneration in Bacillus subtilis. Bradley J...examined in the gram-positive bacterium Bacillus subtilis. Homogenates of this bacterium were able to convert ketomethiobutyrate to methionine, utilising

  15. Ornithine-δ-Aminotransferase Inhibits Neurogenesis During Xenopus Embryonic Development

    PubMed Central

    Peng, Ying; Cooper, Sandra K.; Li, Yi; Mei, Jay M.; Qiu, Shuwei; Borchert, Gregory L.; Donald, Steven P.; Kung, Hsiang-fu; Phang, James M.

    2015-01-01

    Purpose. In humans, deficiency of ornithine-δ-aminotransferase (OAT) results in progressive degeneration of the neural retina (gyrate atrophy) with blindness in the fourth decade. In this study, we used the Xenopus embryonic developmental model to study functions of the OAT gene on embryonic development. Methods. We cloned and sequenced full-length OAT cDNA from Xenopus oocytes (X-OAT) and determined X-OAT expression in various developmental stages of Xenopus embryos and in a variety of adult tissues. The phenotype, gene expression of neural developmental markers, and enzymatic activity were detected by gain-of-function and loss-of-function manipulations. Results. We showed that X-OAT is essential for Xenopus embryonic development, and overexpression of X-OAT produces a ventralized phenotype characterized by a small head, lack of axial structure, and defective expression of neural developmental markers. Using X-OAT mutants based on mutations identified in humans, we found that substitution of both Arg 180 and Leu 402 abrogated both X-OAT enzymatic activity and ability to modulate the developmental phenotype. Neurogenesis is inhibited by X-OAT during Xenopus embryonic development. Conclusions. Neurogenesis is inhibited by X-OAT during Xenopus embryonic development, but it is essential for Xenopus embryonic development. The Arg 180 and Leu 402 are crucial for these effects of the OAT molecule in development. PMID:25783604

  16. Tyrosine aminotransferase from Leishmania infantum: A new drug target candidate

    PubMed Central

    Moreno, Miguel Angel; Alonso, Ana; Alcolea, Pedro Jose; Abramov, Ariel; de Lacoba, Mario García; Abendroth, Jan; Zhang, Sunny; Edwards, Thomas; Lorimer, Don; Myler, Peter John; Larraga, Vicente

    2014-01-01

    Leishmania infantum is the etiological agent of zoonotic visceral leishmaniasis in the Mediterranean basin. The disease is fatal without treatment, which has been based on antimonial pentavalents for more than 60 years. Due to resistances, relapses and toxicity to current treatment, the development of new drugs is required. The structure of the L. infantum tyrosine aminotransferase (LiTAT) has been recently solved showing important differences with the mammalian orthologue. The characterization of LiTAT is reported herein. This enzyme is cytoplasmic and is over-expressed in the more infective stages and nitric oxide resistant parasites. Unlike the mammalian TAT, LiTAT is able to use ketomethiobutyrate as co-substrate. The pharmacophore model of LiTAT with this specific co-substrate is described herein. This may allow the identification of new inhibitors present in the databases. All the data obtained support that LiTAT is a good target candidate for the development of new anti-leishmanial drugs. PMID:25516846

  17. Design and Mechanism of Tetrahydrothiophene-based GABA Aminotransferase Inactivators

    PubMed Central

    Le, Hoang V.; Hawker, Dustin D.; Wu, Rui; Doud, Emma; Widom, Julia; Sanishvili, Ruslan; Liu, Dali; Kelleher, Neil L.; Silverman, Richard B.

    2015-01-01

    Low levels of γ-aminobutyric acid (GABA), one of two major neurotransmitters that regulate brain neuronal activity, are associated with many neurological disorders, such as epilepsy, Parkinson’s disease, Alzheimer’s disease, Huntington’s disease, and cocaine addiction. One of the main methods to raise the GABA level in human brain is to use small molecules that cross the blood-brain barrier and inhibit the activity of γ-aminobutyric acid aminotransferase (GABA-AT), the enzyme that degrades GABA. We have designed a series of conformationally-restricted, tetrahydrothiophene-based GABA analogs with a properly-positioned leaving group that could facilitate a ring-opening mechanism, leading to inactivation of GABA-AT. One compound in the series is eight times more efficient an inactivator of GABA-AT than vigabatrin, the only FDA-approved inactivator of GABA-AT. Our mechanistic studies show that the compound inactivates GABA-AT by a new mechanism. The metabolite resulting from inactivation does not covalently bind to amino acid residues of GABA-AT but stays in the active site via H-bond interactions with Arg-192, a π-π interaction with Phe-189, and a weak nonbonded S···O=C interaction with Glu-270, thereby inactivating the enzyme. PMID:25781189

  18. Cloning and expression of human tyrosine aminotransferase cDNA.

    PubMed

    Séralini, G E; Luu-Thé, V; Labrie, F

    1995-01-02

    Complementary DNA clones encoding human tyrosine aminotransferase (TAT) were isolated by screening a normal adult woman liver lambda gt11 library with rat TAT cDNA. The largest isolated cDNA is 2051 bp long (EMBL accession number X55675). This cDNA was subcloned downstream of the cytomegalovirus promoter in the pCMV vector for transfection into human cervical carcinoma HeLa cells. Expression of the TAT cDNA resulted in the synthesis of a protein with a molecular mass of approximately 50 kDa, as assessed by Western analysis, a value which is in close agreement with the predicted molecular weight of 50,399, for a deduced sequence of 454 amino acids. The expressed protein catalyzed specifically the conversion of L-[14C]tyrosine into p-[14C]hydroxyphenylpyruvate. The availability of a functional TAT cDNA provides a useful tool for detailed study of the structure-function relationship of the enzyme and its mutated derivatives.

  19. Expression and processing of human ornithine-delta-aminotransferase in Saccharomyces cerevisiae.

    PubMed

    Dougherty, K M; Swanson, D A; Brody, L C; Valle, D

    1993-11-01

    Ornithine-delta-aminotransferase catalyzes the conversion of ornithine to glutamate-gamma-semialdehyde. In humans, deficiency of this mitochondrial matrix enzyme results in the progressive blinding disorder, gyrate atrophy of the choroid and retina. To explore yeast as an expression system, we introduced a cDNA encoding human ornithine-delta-aminotransferase into an ornithine aminotransferase-deficient strain of Saccharomyces cerevisiae. The human enzyme was expressed at high levels, with activity 20-fold greater than that of wild-type yeast and 10-fold higher than in human fibroblasts. Although the normal location of ornithine-delta-aminotransferase in S. cerevisiae is cytosolic, human ornithine-delta-aminotransferase expressed in S. cerevisiae was localized to the mitochondrial matrix with correct proteolytic processing of its mitochondrial leader sequence. Despite this anomalous location in yeast, human ornithine-delta-aminotransferase complemented the phenotype of the mutant strain, restoring its ability to utilize ornithine as a sole nitrogen source. We also expressed a vitamin B6-responsive missense allele of ornithine-delta-aminotransferase (V332M) and showed that the biochemical phenotype of this allele is easily demonstrated confirming the usefulness of this system for examining mutations causing gyrate atrophy.

  20. Sensitive non-radioactive determination of aminotransferase stereospecificity for C-4' hydrogen transfer on the coenzyme.

    PubMed

    Jomrit, Juntratip; Summpunn, Pijug; Meevootisom, Vithaya; Wiyakrutta, Suthep

    2011-02-25

    A sensitive non-radioactive method for determination of the stereospecificity of the C-4' hydrogen transfer on the coenzymes (pyridoxal phosphate, PLP; and pyridoxamine phosphate, PMP) of aminotransferases has been developed. Aminotransferase of unknown stereospecificity in its PLP form was incubated in (2)H(2)O with a substrate amino acid resulted in PMP labeled with deuterium at C-4' in the pro-S or pro-R configuration according to the stereospecificity of the aminotransferase tested. The [4'-(2)H]PMP was isolated from the enzyme protein and divided into two portions. The first portion was incubated in aqueous buffer with apo-aspartate aminotransferase (a reference si-face specific enzyme), and the other was incubated with apo-branched-chain amino acid aminotransferase (a reference re-face specific enzyme) in the presence of a substrate 2-oxo acid. The (2)H at C-4' is retained with the PLP if the aminotransferase in question transfers C-4' hydrogen on the opposite face of the coenzyme compared with the reference aminotransferase, but the (2)H is removed if the test and reference aminotransferases catalyze hydrogen transfer on the same face. PLP formed in the final reactions was analyzed by LC-MS/MS for the presence or absence of (2)H. The method was highly sensitive that for the aminotransferase with ca. 50 kDa subunit molecular weight, only 2mg of the enzyme was sufficient for the whole test. With this method, the use of radioactive substances could be avoided without compromising the sensitivity of the assay.

  1. Sensitive non-radioactive determination of aminotransferase stereospecificity for C-4' hydrogen transfer on the coenzyme

    SciTech Connect

    Jomrit, Juntratip; Summpunn, Pijug; Meevootisom, Vithaya; Wiyakrutta, Suthep

    2011-02-25

    Research highlights: {yields} Stereochemical mechanism of PLP enzymes is important but difficult to determine. {yields} This new method is significantly less complicated than the previous ones. {yields} This assay is as sensitive as the radioactive based method. {yields} LC-MS/MS positively identify the analyte coenzyme. {yields} The method can be used with enzyme whose apo form is unstable. -- Abstract: A sensitive non-radioactive method for determination of the stereospecificity of the C-4' hydrogen transfer on the coenzymes (pyridoxal phosphate, PLP; and pyridoxamine phosphate, PMP) of aminotransferases has been developed. Aminotransferase of unknown stereospecificity in its PLP form was incubated in {sup 2}H{sub 2}O with a substrate amino acid resulted in PMP labeled with deuterium at C-4' in the pro-S or pro-R configuration according to the stereospecificity of the aminotransferase tested. The [4'-{sup 2}H]PMP was isolated from the enzyme protein and divided into two portions. The first portion was incubated in aqueous buffer with apo-aspartate aminotransferase (a reference si-face specific enzyme), and the other was incubated with apo-branched-chain amino acid aminotransferase (a reference re-face specific enzyme) in the presence of a substrate 2-oxo acid. The {sup 2}H at C-4' is retained with the PLP if the aminotransferase in question transfers C-4' hydrogen on the opposite face of the coenzyme compared with the reference aminotransferase, but the {sup 2}H is removed if the test and reference aminotransferases catalyze hydrogen transfer on the same face. PLP formed in the final reactions was analyzed by LC-MS/MS for the presence or absence of {sup 2}H. The method was highly sensitive that for the aminotransferase with ca. 50 kDa subunit molecular weight, only 2 mg of the enzyme was sufficient for the whole test. With this method, the use of radioactive substances could be avoided without compromising the sensitivity of the assay.

  2. Serum Alanine Aminotransferase Levels, Hematocrit Rate and Body Weight Correlations Before and After Hemodialysis Session

    PubMed Central

    Lopes, Edmundo Pessoa; Sette, Luis Henrique B. C.; Sette, Jorge Bezerra C.; Luna, Carlos F.; Andrade, Amaro M.; Moraes, Maviael; Sette, Paulo C. A.; Menezes, Roberto; Cavalcanti, Rui L.; Conceição, Sergio C.

    2009-01-01

    PURPOSE To evaluate alanine aminotransferase levels before and after a hemodialysis session and to correlate these values with the hematocrit rate and weight loss during hemodialysis. PATIENTS AND METHODS The serum alanine aminotransferase levels, hematocrit rate and body weight were measured and correlated before and after a single hemodialysis session for 146 patients with chronic renal failure. An receiver operating characteristic (ROC) curve for the serum alanine aminotransferase levels collected before and after hemodialysis was plotted to identify hepatitis C virus-infected patients. RESULTS The mean weight loss of the 146 patients during hemodialysis was 5.3% (p < 0.001). The mean alanine aminotransferase levels before and after hemodialysis were 18.8 and 23.9 IU/, respectively, denoting a significant 28.1% increase. An equally significant increase of 16.4% in the hematocrit rate also occurred after hemodialysis. The weight loss was inversely correlated with the rise in both the alanine aminotransferase level (r = 0.3; p < 0.001) and hematocrit rate (r = 0.5; p < 0.001). A direct correlation was found between the rise in alanine aminotransferase levels and the hematocrit during the hemodialysis session (r = 0.4; p < 0.001). Based on the ROC curve, the upper limit of the normal alanine aminotransferase level should be reduced by 40% relative to the upper limit of normal if the blood samples are collected before the hemodialysis session or by 60% if blood samples are collected after the session. CONCLUSION In the present study, significant elevations in the serum alanine aminotransferase levels and hematocrit rates occurred in parallel to a reduction in body weight after the hemodialysis session. These findings suggest that one of the factors for low alanine aminotransferase levels prior to hemodialysis could be hemodilution in patients with chronic renal failure. PMID:19841699

  3. Alanine Aminotransferase Variants Conferring Diverse NUE Phenotypes in Arabidopsis thaliana

    PubMed Central

    McAllister, Chandra H.; Good, Allen G.

    2015-01-01

    Alanine aminotransferase (AlaAT, E.C. 2.6.1.2), is a pyridoxal-5’-phosphate-dependent (PLP) enzyme that catalyzes the reversible transfer of an amino group from alanine to 2-oxoglutarate to produce glutamate and pyruvate, or vice versa. It has been well documented in both greenhouse and field studies that tissue-specific over-expression of AlaAT from barley (Hordeum vulgare, HvAlaAT) results in a significant increase in plant NUE in both canola and rice. While the physical phenotypes associated with over-expression of HvAlaAT have been well characterized, the role this enzyme plays in vivo to create a more N efficient plant remains unknown. Furthermore, the importance of HvAlaAT, in contrast to other AlaAT enzyme homologues in creating this phenotype has not yet been explored. To address the role of AlaAT in NUE, AlaAT variants from diverse sources and different subcellular locations, were expressed in the wild-type Arabidopsis thaliana Col-0 background and alaat1;2 (alaat1-1;alaat2-1) knockout background in various N environments. The analysis and comparison of both the physical and physiological properties of AlaAT over-expressing transgenic plants demonstrated significant differences between plants expressing the different AlaAT enzymes under different external conditions. This analysis indicates that the over-expression of AlaAT variants other than HvAlaAT in crop plants could further increase the NUE phenotype(s) previously observed. PMID:25830496

  4. Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III

    SciTech Connect

    Han, Q.; Robinson, H; Cai, T; Tagle, D; Li, J

    2009-01-01

    Kynurenine aminotransferase III (KAT III) has been considered to be involved in the production of mammalian brain kynurenic acid (KYNA), which plays an important role in protecting neurons from overstimulation by excitatory neurotransmitters. The enzyme was identified based on its high sequence identity with mammalian KAT I, but its activity toward kynurenine and its structural characteristics have not been established. In this study, the biochemical and structural properties of mouse KAT III (mKAT III) were determined. Specifically, mKAT III cDNA was amplified from a mouse brain cDNA library, and its recombinant protein was expressed in an insect cell protein expression system. We established that mKAT III is able to efficiently catalyze the transamination of kynurenine to KYNA and has optimum activity at relatively basic conditions of around pH 9.0 and at relatively high temperatures of 50 to 60C. In addition, mKAT III is active toward a number of other amino acids. Its activity toward kynurenine is significantly decreased in the presence of methionine, histidine, glutamine, leucine, cysteine, and 3-hydroxykynurenine. Through macromolecular crystallography, we determined the mKAT III crystal structure and its structures in complex with kynurenine and glutamine. Structural analysis revealed the overall architecture of mKAT III and its cofactor binding site and active center residues. This is the first report concerning the biochemical characteristics and crystal structures of KAT III enzymes and provides a basis toward understanding the overall physiological role of mammalian KAT III in vivo and insight into regulating the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  5. Biochemical and structural properties of mouse kynurenine aminotransferase III.

    PubMed

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A; Li, Jianyong

    2009-02-01

    Kynurenine aminotransferase III (KAT III) has been considered to be involved in the production of mammalian brain kynurenic acid (KYNA), which plays an important role in protecting neurons from overstimulation by excitatory neurotransmitters. The enzyme was identified based on its high sequence identity with mammalian KAT I, but its activity toward kynurenine and its structural characteristics have not been established. In this study, the biochemical and structural properties of mouse KAT III (mKAT III) were determined. Specifically, mKAT III cDNA was amplified from a mouse brain cDNA library, and its recombinant protein was expressed in an insect cell protein expression system. We established that mKAT III is able to efficiently catalyze the transamination of kynurenine to KYNA and has optimum activity at relatively basic conditions of around pH 9.0 and at relatively high temperatures of 50 to 60 degrees C. In addition, mKAT III is active toward a number of other amino acids. Its activity toward kynurenine is significantly decreased in the presence of methionine, histidine, glutamine, leucine, cysteine, and 3-hydroxykynurenine. Through macromolecular crystallography, we determined the mKAT III crystal structure and its structures in complex with kynurenine and glutamine. Structural analysis revealed the overall architecture of mKAT III and its cofactor binding site and active center residues. This is the first report concerning the biochemical characteristics and crystal structures of KAT III enzymes and provides a basis toward understanding the overall physiological role of mammalian KAT III in vivo and insight into regulating the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  6. Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III▿

    PubMed Central

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A.; Li, Jianyong

    2009-01-01

    Kynurenine aminotransferase III (KAT III) has been considered to be involved in the production of mammalian brain kynurenic acid (KYNA), which plays an important role in protecting neurons from overstimulation by excitatory neurotransmitters. The enzyme was identified based on its high sequence identity with mammalian KAT I, but its activity toward kynurenine and its structural characteristics have not been established. In this study, the biochemical and structural properties of mouse KAT III (mKAT III) were determined. Specifically, mKAT III cDNA was amplified from a mouse brain cDNA library, and its recombinant protein was expressed in an insect cell protein expression system. We established that mKAT III is able to efficiently catalyze the transamination of kynurenine to KYNA and has optimum activity at relatively basic conditions of around pH 9.0 and at relatively high temperatures of 50 to 60°C. In addition, mKAT III is active toward a number of other amino acids. Its activity toward kynurenine is significantly decreased in the presence of methionine, histidine, glutamine, leucine, cysteine, and 3-hydroxykynurenine. Through macromolecular crystallography, we determined the mKAT III crystal structure and its structures in complex with kynurenine and glutamine. Structural analysis revealed the overall architecture of mKAT III and its cofactor binding site and active center residues. This is the first report concerning the biochemical characteristics and crystal structures of KAT III enzymes and provides a basis toward understanding the overall physiological role of mammalian KAT III in vivo and insight into regulating the levels of endogenous KYNA through modulation of the enzyme in the mouse brain. PMID:19029248

  7. Alanine aminotransferase variants conferring diverse NUE phenotypes in Arabidopsis thaliana.

    PubMed

    McAllister, Chandra H; Good, Allen G

    2015-01-01

    Alanine aminotransferase (AlaAT, E.C. 2.6.1.2), is a pyridoxal-5'-phosphate-dependent (PLP) enzyme that catalyzes the reversible transfer of an amino group from alanine to 2-oxoglutarate to produce glutamate and pyruvate, or vice versa. It has been well documented in both greenhouse and field studies that tissue-specific over-expression of AlaAT from barley (Hordeum vulgare, HvAlaAT) results in a significant increase in plant NUE in both canola and rice. While the physical phenotypes associated with over-expression of HvAlaAT have been well characterized, the role this enzyme plays in vivo to create a more N efficient plant remains unknown. Furthermore, the importance of HvAlaAT, in contrast to other AlaAT enzyme homologues in creating this phenotype has not yet been explored. To address the role of AlaAT in NUE, AlaAT variants from diverse sources and different subcellular locations, were expressed in the wild-type Arabidopsis thaliana Col-0 background and alaat1;2 (alaat1-1;alaat2-1) knockout background in various N environments. The analysis and comparison of both the physical and physiological properties of AlaAT over-expressing transgenic plants demonstrated significant differences between plants expressing the different AlaAT enzymes under different external conditions. This analysis indicates that the over-expression of AlaAT variants other than HvAlaAT in crop plants could further increase the NUE phenotype(s) previously observed.

  8. Tyrosine aminotransferase: biochemical and structural properties and molecular dynamics simulations

    SciTech Connect

    Mehere, P.; Robinson, H.; Han, Q.; Lemkul, J. A.; Vavricka, C. J.; Bevan, D. R.; Li, J.

    2010-11-01

    Tyrosine aminotransferase (TAT) catalyzes the transamination of tyrosine and other aromatic amino acids. The enzyme is thought to play a role in tyrosinemia type II, hepatitis and hepatic carcinoma recovery. The objective of this study is to investigate its biochemical and structural characteristics and substrate specificity in order to provide insight regarding its involvement in these diseases. Mouse TAT (mTAT) was cloned from a mouse cDNA library, and its recombinant protein was produced using Escherichia coli cells and purified using various chromatographic techniques. The recombinant mTAT is able to catalyze the transamination of tyrosine using {alpha}-ketoglutaric acid as an amino group acceptor at neutral pH. The enzyme also can use glutamate and phenylalanine as amino group donors and p-hydroxy-phenylpyruvate, phenylpyruvate and alpha-ketocaproic acid as amino group acceptors. Through macromolecular crystallography we have determined the mTAT crystal structure at 2.9 {angstrom} resolution. The crystal structure revealed the interaction between the pyridoxal-5'-phosphate cofactor and the enzyme, as well as the formation of a disulphide bond. The detection of disulphide bond provides some rational explanation regarding previously observed TAT inactivation under oxidative conditions and reactivation of the inactive TAT in the presence of a reducing agent. Molecular dynamics simulations using the crystal structures of Trypanosoma cruzi TAT and human TAT provided further insight regarding the substrate-enzyme interactions and substrate specificity. The biochemical and structural properties of TAT and the binding of its cofactor and the substrate may help in elucidation of the mechanism of TAT inhibition and activation.

  9. Tyrosine Aminotransferase: Biochemical and Structural Properties and Molecular Dynamics Simulations

    SciTech Connect

    P Mehere; Q Han; J Lemkul; C Vavricka; H Robinson; D Bevan; J Li

    2011-12-31

    Tyrosine aminotransferase (TAT) catalyzes the transamination of tyrosine and other aromatic amino acids. The enzyme is thought to play a role in tyrosinemia type II, hepatitis and hepatic carcinoma recovery. The objective of this study is to investigate its biochemical and structural characteristics and substrate specificity in order to provide insight regarding its involvement in these diseases. Mouse TAT (mTAT) was cloned from a mouse cDNA library, and its recombinant protein was produced using Escherichia coli cells and purified using various chromatographic techniques. The recombinant mTAT is able to catalyze the transamination of tyrosine using {alpha}-ketoglutaric acid as an amino group acceptor at neutral pH. The enzyme also can use glutamate and phenylalanine as amino group donors and p-hydroxy-phenylpyruvate, phenylpyruvate and alpha-ketocaproic acid as amino group acceptors. Through macromolecular crystallography we have determined the mTAT crystal structure at 2.9 {angstrom} resolution. The crystal structure revealed the interaction between the pyridoxal-5'-phosphate cofactor and the enzyme, as well as the formation of a disulphide bond. The detection of disulphide bond provides some rational explanation regarding previously observed TAT inactivation under oxidative conditions and reactivation of the inactive TAT in the presence of a reducing agent. Molecular dynamics simulations using the crystal structures of Trypanosoma cruzi TAT and human TAT provided further insight regarding the substrate-enzyme interactions and substrate specificity. The biochemical and structural properties of TAT and the binding of its cofactor and the substrate may help in elucidation of the mechanism of TAT inhibition and activation.

  10. Tyrosine aminotransferase contributes to benzylisoquinoline alkaloid biosynthesis in opium poppy.

    PubMed

    Lee, Eun-Jeong; Facchini, Peter J

    2011-11-01

    Tyrosine aminotransferase (TyrAT) catalyzes the transamination of L-Tyr and α-ketoglutarate, yielding 4-hydroxyphenylpyruvic acid and L-glutamate. The decarboxylation product of 4-hydroxyphenylpyruvic acid, 4-hydroxyphenylacetaldehyde, is a precursor to a large and diverse group of natural products known collectively as benzylisoquinoline alkaloids (BIAs). We have isolated and characterized a TyrAT cDNA from opium poppy (Papaver somniferum), which remains the only commercial source for several pharmaceutical BIAs, including codeine, morphine, and noscapine. TyrAT belongs to group I pyridoxal 5'-phosphate (PLP)-dependent enzymes wherein Schiff base formation occurs between PLP and a specific Lys residue. The amino acid sequence of TyrAT showed considerable homology to other putative plant TyrATs, although few of these have been functionally characterized. Purified, recombinant TyrAT displayed a molecular mass of approximately 46 kD and a substrate preference for L-Tyr and α-ketoglutarate, with apparent K(m) values of 1.82 and 0.35 mm, respectively. No specific requirement for PLP was detected in vitro. Liquid chromatography-tandem mass spectrometry confirmed the conversion of L-Tyr to 4-hydroxyphenylpyruvate. TyrAT gene transcripts were most abundant in roots and stems of mature opium poppy plants. Virus-induced gene silencing was used to evaluate the contribution of TyrAT to BIA metabolism in opium poppy. TyrAT transcript levels were reduced by at least 80% in silenced plants compared with controls and showed a moderate reduction in total alkaloid content. The modest correlation between transcript levels and BIA accumulation in opium poppy supports a role for TyrAT in the generation of alkaloid precursors, but it also suggests the occurrence of other sources for 4-hydroxyphenylacetaldehyde.

  11. Purification and properties of 7, 8-diaminopelargonic acid aminotransferase.

    PubMed

    Stoner, G L; Eisenberg, M A

    1975-06-10

    The enzyme 7, 8-diaminopelargonic acid aminotransferase utilizes S-adenosyl-L-methionine to transaminate the biotin precurson 7-keto-8-aminopelargonic acid and form the next intermediate in the pathway, 7, 8-diaminopelargonic acid. The enzyme has been purified nearly 1000-fold from an extract of a regulatory mutant of Escherichia coli which is derepressed for the enzymes of the biotin operon. The extract was treated with protamine sulfate, ammonium sulfate, and subjected to acid and heat treatments. Subsequently, the enzyme was chromatographed on columns of DEAE-cellulose, phosphocellulose, hydroxylapatite, and two Sephadex G-100. The resulting purified preparation was judged 86% homogeneous by the scanning of of a stained disc gel. The enzymatic activity was associated with the major band in gels run at two different gel concentrations and two different pH values. The cofactor, pyridoxal phosphate, can be resolved from the enzyme in the presence of phosphate buffer after incubation with the amino donor, S-adenosyl-L-methionine. A molecular weight estimation of 94,000 plus or minus 10, 000 has been obtained by gel filtration and sucrose gradient sedimentation studies. Gel electrophoresis in the presence of sodium dodecyl sulfate, shows a single subunit with a molecular weight of 47, 000 plus or minus 3, 000 indicating a dimeric enzyme. A neutral compound was detected in the acidified reaction mixture which was derived from the methionine moiety of S-adenosyl-L-methionine and was present in amounts equivalent to the 7, 8-diaminopelargonic acid produced in the reaction mixture. It is suggested that the keto product of the reaction, i.e. S-adenosyl-2-oxo-4-methylthiobutyric acid, may decompose nonenzymatically under the conditions of the reaction to form 5'-methylthioadenosine and the neutral compound, 2-oxo-3-butenoic acid.

  12. Chronotypic induction of tyrosine aminotransferase by. cap alpha. -methyl-p-tyrosine. [Rat liver

    SciTech Connect

    Cahill, A.L.; Ferguson, S.M.; Ehret, C.F.

    1981-04-06

    ..cap alpha..Methyl-p-tyrosine induced hepatic tyrosine aminotransferase activity to different extents depending upon the time of day of administration of the drug. Maximal induction occurred when ..cap alpha..-methyl-p-tyrosine (100 mg/kg) was injected intraperitoneally during the first several hours of the light phase of the daily cycle, but the magnitude of the induction depended on the nutritional state of the animal. Induction was 4- to 5-fold greater in fasting rats. The effect of ..cap alpha..-methyl-ptyrosine on hepatic tyrosine aminotransferase is believed to be mediated by decreases in hypothalamic norepinephrine. This hypothesis was supported by the demonstration that decreasing levels of hypothalamic norepinephrine at times of day when hypothalamic turnover of norepinephrine was greatest resulted in the greatest induction of tyrosine aminotransferase, while lowering hypothalamic norepinephrine at times when turnover was minimal resulted in minimal induction of tyrosine aminotransferase.

  13. An alternate method for demonstration of erythrocytic aminotransferases on starch gels

    PubMed Central

    Scott, Edward M.; Wright, Rita C.

    1981-01-01

    A two-stage procedure using MTT tetrazolium for the demonstration of aminotransferases (GPT and GOT) either singly or together was developed. Identification of phenotypes was unequivocal in over 99% of the individuals studied. ImagesFig. 1 PMID:7258187

  14. Plastidic aspartate aminotransferases and the biosynthesis of essential amino acids in plants.

    PubMed

    de la Torre, Fernando; Cañas, Rafael A; Pascual, M Belén; Avila, Concepción; Cánovas, Francisco M

    2014-10-01

    In the chloroplasts and in non-green plastids of plants, aspartate is the precursor for the biosynthesis of different amino acids and derived metabolites that play distinct and important roles in plant growth, reproduction, development or defence. Aspartate biosynthesis is mediated by the enzyme aspartate aminotransferase (EC 2.6.1.1), which catalyses the reversible transamination between glutamate and oxaloacetate to generate aspartate and 2-oxoglutarate. Plastids contain two aspartate aminotransferases: a eukaryotic-type and a prokaryotic-type bifunctional enzyme displaying aspartate and prephenate aminotransferase activities. A general overview of the biochemistry, regulation, functional significance, and phylogenetic origin of both enzymes is presented. The roles of these plastidic aminotransferases in the biosynthesis of essential amino acids are discussed.

  15. Biochemical properties and crystal structure of a β-phenylalanine aminotransferase from Variovorax paradoxus.

    PubMed

    Crismaru, Ciprian G; Wybenga, Gjalt G; Szymanski, Wiktor; Wijma, Hein J; Wu, Bian; Bartsch, Sebastian; de Wildeman, Stefaan; Poelarends, Gerrit J; Feringa, Ben L; Dijkstra, Bauke W; Janssen, Dick B

    2013-01-01

    By selective enrichment, we isolated a bacterium that can use β-phenylalanine as a sole nitrogen source. It was identified by 16S rRNA gene sequencing as a strain of Variovorax paradoxus. Enzyme assays revealed an aminotransferase activity. Partial genome sequencing and screening of a cosmid DNA library resulted in the identification of a 1,302-bp aminotransferase gene, which encodes a 46,416-Da protein. The gene was cloned and overexpressed in Escherichia coli. The recombinant enzyme was purified and showed a specific activity of 17.5 U mg(-1) for (S)-β-phenylalanine at 30°C and 33 U mg(-1) at the optimum temperature of 55°C. The β-specific aminotransferase exhibits a broad substrate range, accepting ortho-, meta-, and para-substituted β-phenylalanine derivatives as amino donors and 2-oxoglutarate and pyruvate as amino acceptors. The enzyme is highly enantioselective toward (S)-β-phenylalanine (enantioselectivity [E], >100) and derivatives thereof with different substituents on the phenyl ring, allowing the kinetic resolution of various racemic β-amino acids to yield (R)-β-amino acids with >95% enantiomeric excess (ee). The crystal structures of the holoenzyme and of the enzyme in complex with the inhibitor 2-aminooxyacetate revealed structural similarity to the β-phenylalanine aminotransferase from Mesorhizobium sp. strain LUK. The crystal structure was used to rationalize the stereo- and regioselectivity of V. paradoxus aminotransferase and to define a sequence motif with which new aromatic β-amino acid-converting aminotransferases may be identified.

  16. Clinicopathological features of choledocholithiasis patients with high aminotransferase levels without cholangitis

    PubMed Central

    Huh, Cheal Wung; Jang, Sung Ill; Lim, Beom Jin; Kim, Hee Wook; Kim, Jae Keun; Park, Jun Sung; Kim, Ja Kyung; Lee, Se Joon; Lee, Dong Ki

    2016-01-01

    Abstract Common bile duct (CBD) stones are generally associated with greater elevations of alkaline phosphatase and gamma-glutamyl transpeptidase levels than aspartate aminotransferase and alanine aminotransferase levels. However, some patients with CBD stones show markedly increased aminotransferase levels, sometimes leading to the misdiagnosis of liver disease. Therefore, the aim of this study was to investigate the clinicopathologic features of patients with CBD stones and high aminotransferase levels. This prospective cohort study included 882 patients diagnosed with CBD stones using endoscopic retrograde cholangiopancreatography (ERCP). Among these patients, 38 (4.3%) exhibited aminotransferase levels above 400 IU/L without cholangitis (gallstone hepatitis [GSH] group), and 116 (13.2%) exhibited normal aminotransferase levels (control group). We compared groups in terms of clinical features, laboratory test results, radiologic images, and ERCP findings such as CBD diameter, CBD stone diameter and number, and periampullary diverticulum. Liver biopsy was performed for patients in the GSH group. GSH patients were younger and more likely to have gallbladder stones than control patients, implying a higher incidence of gallbladder stone migration. Also, GSH patients experienced more severe, short-lasting abdominal pain. ERCP showed narrower CBDs in GSH patients than in control patients. Histological analysis of liver tissue from GSH patients showed no abnormalities except for mild inflammation. Compared with control patients, GSH patients were younger and showed more severe, short-lasting abdominal pain, which could be due to a sudden increase of CBD pressure resulting from the migration of gallstones through narrower CBDs. These clinical features could be helpful not only for the differential diagnosis of liver disease but also for investigating the underlying mechanisms of liver damage in obstructive jaundice. Moreover, we propose a new definition of

  17. Biochemical and Structural Characterization of a Ureidoglycine Aminotransferase in the Klebsiella pneumoniae Uric Acid Catabolic Pathway

    SciTech Connect

    French, Jarrod B.; Ealick, Steven E.

    2010-09-03

    Many plants, fungi, and bacteria catabolize allantoin as a mechanism for nitrogen assimilation. Recent reports have shown that in plants and some bacteria the product of hydrolysis of allantoin by allantoinase is the unstable intermediate ureidoglycine. While this molecule can spontaneously decay, genetic analysis of some bacterial genomes indicates that an aminotransferase may be present in the pathway. Here we present evidence that Klebsiella pneumoniae HpxJ is an aminotransferase that preferentially converts ureidoglycine and an {alpha}-keto acid into oxalurate and the corresponding amino acid. We determined the crystal structure of HpxJ, allowing us to present an explanation for substrate specificity.

  18. Beta-alanine/alpha-ketoglutarate aminotransferase for 3-hydroxypropionic acid production

    DOEpatents

    Jessen, Holly Jean [Chanhassen, MN; Liao, Hans H [Eden Prairie, MN; Gort, Steven John [Apple Valley, MN; Selifonova, Olga V [Plymouth, MN

    2011-10-04

    The present disclosure provides novel beta-alanine/alpha ketoglutarate aminotransferase nucleic acid and protein sequences having increased biological activity. Also provided are cells containing such enzymes, as well as methods of their use, for example to produce malonyl semialdehyde and downstream products thereof, such as 3-hydroxypropionic acid and derivatives thereof.

  19. Structure, expression, and function of kynurenine aminotransferases in human and rodent brains.

    PubMed

    Han, Qian; Cai, Tao; Tagle, Danilo A; Li, Jianyong

    2010-02-01

    Kynurenine aminotransferases (KATs) catalyze the synthesis of kynurenic acid (KYNA), an endogenous antagonist of N-methyl-D: -aspartate and alpha 7-nicotinic acetylcholine receptors. Abnormal KYNA levels in human brains are implicated in the pathophysiology of schizophrenia, Alzheimer's disease, and other neurological disorders. Four KATs have been reported in mammalian brains, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase. KAT II has a striking tertiary structure in N-terminal part and forms a new subgroup in fold type I aminotransferases, which has been classified as subgroup Iepsilon. Knowledge regarding KATs is vast and complex; therefore, this review is focused on recent important progress of their gene characterization, physiological and biochemical function, and structural properties. The biochemical differences of four KATs, specific enzyme activity assays, and the structural insights into the mechanism of catalysis and inhibition of these enzymes are discussed.

  20. Structure, expression, and function of kynurenine aminotransferases in human and rodent brains

    PubMed Central

    Han, Qian; Cai, Tao; Tagle, Danilo A.

    2010-01-01

    Kynurenine aminotransferases (KATs) catalyze the synthesis of kynurenic acid (KYNA), an endogenous antagonist of N-methyl-d-aspartate and alpha 7-nicotinic acetylcholine receptors. Abnormal KYNA levels in human brains are implicated in the pathophysiology of schizophrenia, Alzheimer's disease, and other neurological disorders. Four KATs have been reported in mammalian brains, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase. KAT II has a striking tertiary structure in N-terminal part and forms a new subgroup in fold type I aminotransferases, which has been classified as subgroup Iε. Knowledge regarding KATs is vast and complex; therefore, this review is focused on recent important progress of their gene characterization, physiological and biochemical function, and structural properties. The biochemical differences of four KATs, specific enzyme activity assays, and the structural insights into the mechanism of catalysis and inhibition of these enzymes are discussed. PMID:19826765

  1. Structural Insights into a Novel Class of Aspartate Aminotransferase from Corynebacterium glutamicum

    PubMed Central

    Son, Hyeoncheol Francis; Kim, Kyung-Jin

    2016-01-01

    Aspartate aminotransferase from Corynebacterium glutamicum (CgAspAT) is a PLP-dependent enzyme that catalyzes the production of L-aspartate and α-ketoglutarate from L-glutamate and oxaloacetate in L-lysine biosynthesis. In order to understand the molecular mechanism of CgAspAT and compare it with those of other aspartate aminotransferases (AspATs) from the aminotransferase class I, we determined the crystal structure of CgAspAT. CgAspAT functions as a dimer, and the CgAspAT monomer consists of two domains, the core domain and the auxiliary domain. The PLP cofactor is found to be bound to CgAspAT and stabilized through unique residues. In our current structure, a citrate molecule is bound at the active site of one molecule and mimics binding of the glutamate substrate. The residues involved in binding of the PLP cofactor and the glutamate substrate were confirmed by site-directed mutagenesis. Interestingly, compared with other AspATs from aminotransferase subgroup Ia and Ib, CgAspAT exhibited unique binding sites for both cofactor and substrate; moreover, it was found to have unusual structural features in the auxiliary domain. Based on these structural differences, we propose that CgAspAT does not belong to either subgroup Ia or Ib, and can be categorized into a subgroup Ic. The phylogenetic tree and RMSD analysis also indicates that CgAspAT is located in an independent AspAT subgroup. PMID:27355211

  2. Structure Expression and Function of kynurenine Aminotransferases in Human and Rodent Brains

    SciTech Connect

    Q Han; T Cai; D Tagle; J Li

    2011-12-31

    Kynurenine aminotransferases (KATs) catalyze the synthesis of kynurenic acid (KYNA), an endogenous antagonist of N-methyl-D: -aspartate and alpha 7-nicotinic acetylcholine receptors. Abnormal KYNA levels in human brains are implicated in the pathophysiology of schizophrenia, Alzheimer's disease, and other neurological disorders. Four KATs have been reported in mammalian brains, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase. KAT II has a striking tertiary structure in N-terminal part and forms a new subgroup in fold type I aminotransferases, which has been classified as subgroup Iepsilon. Knowledge regarding KATs is vast and complex; therefore, this review is focused on recent important progress of their gene characterization, physiological and biochemical function, and structural properties. The biochemical differences of four KATs, specific enzyme activity assays, and the structural insights into the mechanism of catalysis and inhibition of these enzymes are discussed.

  3. PPAR{alpha} regulates the hepatotoxic biomarker alanine aminotransferase (ALT1) gene expression in human hepatocytes

    SciTech Connect

    Thulin, Petra; Rafter, Ingalill; Stockling, Kenneth; Tomkiewicz, Celine; Norjavaara, Ensio; Aggerbeck, Martine; Hellmold, Heike; Ehrenborg, Ewa; Andersson, Ulf; Cotgreave, Ian; Glinghammar, Bjoern

    2008-08-15

    In this work, we investigated a potential mechanism behind the observation of increased aminotransferase levels in a phase I clinical trial using a lipid-lowering drug, the peroxisome proliferator-activated receptor (PPAR) {alpha} agonist, AZD4619. In healthy volunteers treated with AZD4619, serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities were elevated without an increase in other markers for liver injury. These increases in serum aminotransferases have previously been reported in some patients receiving another PPAR{alpha} agonist, fenofibrate. In subsequent in vitro studies, we observed increased expression of ALT1 protein and mRNA in human hepatocytes after treatment with fenofibric acid. The PPAR effect on ALT1 expression was shown to act through a direct transcriptional mechanism involving at least one PPAR response element (PPRE) in the proximal ALT1 promoter, while no effect of fenofibrate and AZD4619 was observed on the ALT2 promoter. Binding of PPARs to the PPRE located at - 574 bp from the transcriptional start site was confirmed on both synthetic oligonucleotides and DNA in hepatocytes. These data show that intracellular ALT expression is regulated by PPAR agonists and that this mechanism might contribute to increased ALT activity in serum.

  4. Structural studies of Pseudomonas and Chromobacterium ω-aminotransferases provide insights into their differing substrate specificity

    SciTech Connect

    Sayer, Christopher; Isupov, Michail N.; Westlake, Aaron; Littlechild, Jennifer A.

    2013-04-01

    The X-ray structures of two ω-aminotransferases from P. aeruginosa and C. violaceum in complex with an inhibitor offer the first detailed insight into the structural basis of the substrate specificity of these industrially important enzymes. The crystal structures and inhibitor complexes of two industrially important ω-aminotransferase enzymes from Pseudomonas aeruginosa and Chromobacterium violaceum have been determined in order to understand the differences in their substrate specificity. The two enzymes share 30% sequence identity and use the same amino acceptor, pyruvate; however, the Pseudomonas enzyme shows activity towards the amino donor β-alanine, whilst the Chromobacterium enzyme does not. Both enzymes show activity towards S-α-methylbenzylamine (MBA), with the Chromobacterium enzyme having a broader substrate range. The crystal structure of the P. aeruginosa enzyme has been solved in the holo form and with the inhibitor gabaculine bound. The C. violaceum enzyme has been solved in the apo and holo forms and with gabaculine bound. The structures of the holo forms of both enzymes are quite similar. There is little conformational difference observed between the inhibitor complex and the holoenzyme for the P. aeruginosa aminotransferase. In comparison, the crystal structure of the C. violaceum gabaculine complex shows significant structural rearrangements from the structures of both the apo and holo forms of the enzyme. It appears that the different rigidity of the protein scaffold contributes to the substrate specificity observed for the two ω-aminotransferases.

  5. Beta-alanine/alpha-ketoglutarate aminotransferase for 3-hydroxypropionic acid production

    SciTech Connect

    Jessen, Holly Jean; Liao, Hans H; Gort, Steven John; Selifonova, Olga V

    2014-11-18

    The present disclosure provides novel beta-alanine/alpha ketoglutarate aminotransferase nucleic acid and protein sequences having increased biological activity. Also provided are cells containing such enzymes, as well as methods of their use, for example to produce malonyl semialdehyde and downstream products thereof, such as 3-hydroxypropionic acid and derivatives thereof.

  6. Studies on some kinetic parameters of aminotransferases in tissues of the snail, Pila globosa (Swainson) during malathion intoxication.

    PubMed

    Sahib, I K; Rao, K R

    1988-01-01

    The aspartate and alanine aminotransferases in the tissues of the snail, Pila globosa showed high catalytic potentials (low Km and high Vmax) during malathion exposure in vivo. In vitro addition of different concentrations of malathion did not influence aminotransferase activity. The results are discussed in relation to the regulative influence of the intracellular environment of the cell.

  7. Crystal structure of Trypanosoma cruzi tyrosine aminotransferase: substrate specificity is influenced by cofactor binding mode.

    PubMed Central

    Blankenfeldt, W.; Nowicki, C.; Montemartini-Kalisz, M.; Kalisz, H. M.; Hecht, H. J.

    1999-01-01

    The crystal structure of tyrosine aminotransferase (TAT) from the parasitic protozoan Trypanosoma cruzi, which belongs to the aminotransferase subfamily Igamma, has been determined at 2.5 A resolution with the R-value R = 15.1%. T. cruzi TAT shares less than 15% sequence identity with aminotransferases of subfamily Ialpha but shows only two larger topological differences to the aspartate aminotransferases (AspATs). First, TAT contains a loop protruding from the enzyme surface in the larger cofactor-binding domain, where the AspATs have a kinked alpha-helix. Second, in the smaller substrate-binding domain, TAT has a four-stranded antiparallel beta-sheet instead of the two-stranded beta-sheet in the AspATs. The position of the aromatic ring of the pyridoxal-5'-phosphate cofactor is very similar to the AspATs but the phosphate group, in contrast, is closer to the substrate-binding site with one of its oxygen atoms pointing toward the substrate. Differences in substrate specificities of T. cruzi TAT and subfamily Ialpha aminotransferases can be attributed by modeling of substrate complexes mainly to this different position of the cofactor-phosphate group. Absence of the arginine, which in the AspATs fixes the substrate side-chain carboxylate group by a salt bridge, contributes to the inability of T. cruzi TAT to transaminate acidic amino acids. The preference of TAT for tyrosine is probably related to the ability of Asn17 in TAT to form a hydrogen bond to the tyrosine side-chain hydroxyl group. PMID:10595543

  8. Isolation and characterization of a gene coding for a novel aspartate aminotransferase from Rhizobium meliloti.

    PubMed Central

    Alfano, J R; Kahn, M L

    1993-01-01

    Aspartate aminotransferase (AAT) is an important enzyme in aspartate catabolism and biosynthesis and, by converting tricarboxylic acid cycle intermediates to amino acids, AAT is also significant in linking carbon metabolism with nitrogen metabolism. To examine the role of AAT in symbiotic nitrogen fixation further, plasmids encoding three different aminotransferases from Rhizobium meliloti 104A14 were isolated by complementation of an Escherichia coli auxotroph that lacks three aminotransferases. pJA10 contained a gene, aatB, that coded for a previously undescribed AAT, AatB. pJA30 encoded an aromatic aminotransferase, TatA, that had significant AAT activity, and pJA20 encoded a branched-chain aminotransferase designated BatA. Genes for the latter two enzymes, tatA and batA, were previously isolated from R. meliloti. aatB is distinct from but hybridizes to aatA, which codes for AatA, a protein required for symbiotic nitrogen fixation. The DNA sequence of aatB contained an open reading frame that could encode a protein 410 amino acids long and with a monomer molecular mass of 45,100 Da. The amino acid sequence of aatB is unusual, and AatB appears to be a member of a newly described class of AATs. AatB expressed in E. coli has a Km for aspartate of 5.3 mM and a Km for 2-oxoglutarate of 0.87 mM. Its pH optimum is between 8.0 and 8.5. Mutations were constructed in aatB and tatA and transferred to the genome of R. meliloti 104A14. Both mutants were prototrophs and were able to carry out symbiotic nitrogen fixation. Images PMID:8320232

  9. Participation of cysteine and cystine in inactivation of tyrosine aminotransferase in rat liver homogenates.

    PubMed Central

    Buckley, W T; Milligan, L P

    1978-01-01

    1. Inactivation of tyrosine aminotransferase was studied in rat liver homogenates. Under an O2 atmosphere with cysteine added, inactivation was rapid after a lag period of approx. 1h, whereas a N2 atmosphere extended the lag period to approx. 3h. 2. Replacement of cysteine with cystine resulted in rapid inactivation both aerobically and anaerobically. 3. Removal of the particulate fraction by centrifuging rat liver homogenates at 13,000g for 9min resulted in an aerobic lag period of 0.5h in the presence of cystine and approx. 3h in the presence of cysteine. 4. It is proposed that the stimulatory effect of cysteine on tyrosine aminotransferase inactivation occurs largely as a result of oxidation to cystine, which appears to be a more directly effective agent. PMID:33669

  10. Deciphering the role of aspartate and prephenate aminotransferase activities in plastid nitrogen metabolism.

    PubMed

    de la Torre, Fernando; El-Azaz, Jorge; Avila, Concepción; Cánovas, Francisco M

    2014-01-01

    Chloroplasts and plastids of nonphotosynthetic plant cells contain two aspartate (Asp) aminotransferases: a eukaryotic type (Asp5) and a prokaryotic-type bifunctional enzyme displaying Asp and prephenate aminotransferase activities (PAT). We have identified the entire Asp aminotransferase gene family in Nicotiana benthamiana and isolated and cloned the genes encoding the isoenzymes with plastidic localization: NbAsp5 and NbPAT. Using a virus-induced gene silencing approach, we obtained N. benthamiana plants silenced for NbAsp5 and/or NbPAT. Phenotypic and metabolic analyses were conducted in silenced plants to investigate the specific roles of these enzymes in the biosynthesis of essential amino acids within the plastid. The NbAsp5 silenced plants had no changes in phenotype, exhibiting similar levels of free Asp and glutamate as control plants, but contained diminished levels of asparagine and much higher levels of lysine. In contrast, the suppression of NbPAT led to a severe reduction in growth and strong chlorosis symptoms. NbPAT silenced plants exhibited extremely reduced levels of asparagine and were greatly affected in their phenylalanine metabolism and lignin deposition. Furthermore, NbPAT suppression triggered a transcriptional reprogramming in plastid nitrogen metabolism. Taken together, our results indicate that NbPAT has an overlapping role with NbAsp5 in the biosynthesis of Asp and a key role in the production of phenylalanine for the biosynthesis of phenylpropanoids. The analysis of NbAsp5/NbPAT cosilenced plants highlights the central role of both plastidic aminotransferases in nitrogen metabolism; however, only NbPAT is essential for plant growth and development.

  11. Identification and expression analyses of the alanine aminotransferase (AlaAT) gene family in poplar seedlings.

    PubMed

    Xu, Zhiru; Ma, Jing; Qu, Chunpu; Hu, Yanbo; Hao, Bingqing; Sun, Yan; Liu, Zhongye; Yang, Han; Yang, Chengjun; Wang, Hongwei; Li, Ying; Liu, Guanjun

    2017-04-05

    Alanine aminotransferase (AlaAT, E.C.2.6.1.2) catalyzes the reversible conversion of pyruvate and glutamate to alanine and α-oxoglutarate. The AlaAT gene family has been well studied in some herbaceous plants, but has not been well characterized in woody plants. In this study, we identified four alanine aminotransferase homologues in Populus trichocarpa, which could be classified into two subgroups, A and B. AlaAT3 and AlaAT4 in subgroup A encode AlaAT, while AlaAT1 and AlaAT2 in subgroup B encode glutamate:glyoxylate aminotransferase (GGAT), which catalyzes the reaction of glutamate and glyoxylate to α-oxoglutarate and glycine. Four AlaAT genes were cloned from P. simonii × P. nigra. PnAlaAT1 and PnAlaAT2 were expressed predominantly in leaves and induced by exogenous nitrogen and exhibited a diurnal fluctuation in leaves, but was inhibited in roots. PnAlaAT3 and PnAlaAT4 were mainly expressed in roots, stems and leaves, and was induced by exogenous nitrogen. The expression of PnAlaAT3 gene could be regulated by glutamine or its related metabolites in roots. Our results suggest that PnAlaAT3 gene may play an important role in nitrogen metabolism and is regulated by glutamine or its related metabolites in the roots of P. simonii × P. nigra.

  12. Protein Homeostasis Defects of Alanine-Glyoxylate Aminotransferase: New Therapeutic Strategies in Primary Hyperoxaluria Type I

    PubMed Central

    Pey, Angel L.; Albert, Armando; Salido, Eduardo

    2013-01-01

    Alanine-glyoxylate aminotransferase catalyzes the transamination between L-alanine and glyoxylate to produce pyruvate and glycine using pyridoxal 5′-phosphate (PLP) as cofactor. Human alanine-glyoxylate aminotransferase is a peroxisomal enzyme expressed in the hepatocytes, the main site of glyoxylate detoxification. Its deficit causes primary hyperoxaluria type I, a rare but severe inborn error of metabolism. Single amino acid changes are the main type of mutation causing this disease, and considerable effort has been dedicated to the understanding of the molecular consequences of such missense mutations. In this review, we summarize the role of protein homeostasis in the basic mechanisms of primary hyperoxaluria. Intrinsic physicochemical properties of polypeptide chains such as thermodynamic stability, folding, unfolding, and misfolding rates as well as the interaction of different folding states with protein homeostasis networks are essential to understand this disease. The view presented has important implications for the development of new therapeutic strategies based on targeting specific elements of alanine-glyoxylate aminotransferase homeostasis. PMID:23956997

  13. Weaning Induced Hepatic Oxidative Stress, Apoptosis, and Aminotransferases through MAPK Signaling Pathways in Piglets

    PubMed Central

    Luo, Zhen; Zhu, Wei; Guo, Qi; Luo, Wenli; Zhang, Jing; Xu, Weina

    2016-01-01

    This study investigated the effects of weaning on the hepatic redox status, apoptosis, function, and the mitogen-activated protein kinase (MAPK) signaling pathways during the first week after weaning in piglets. A total of 12 litters of piglets were weaned at d 21 and divided into the weaning group (WG) and the control group (CG). Six piglets from each group were slaughtered at d 0 (d 20, referred to weaning), d 1, d 4, and d 7 after weaning. Results showed that weaning significantly increased the concentrations of hepatic free radicals H2O2 and NO, malondialdehyde (MDA), and 8-hydroxy-2′-deoxyguanosine (8-OHdG), while significantly decreasing the inhibitory hydroxyl ability (IHA) and glutathione peroxidase (GSH-Px), and altered the level of superoxide dismutase (SOD). The apoptosis results showed that weaning increased the concentrations of caspase-3, caspase-8, caspase-9 and the ratio of Bax/Bcl-2. In addition, aspartate aminotransferase transaminase (AST) and alanine aminotransferase (ALT) in liver homogenates increased after weaning. The phosphorylated JNK and ERK1/2 increased, while the activated p38 initially decreased and then increased. Our results suggested that weaning increased the hepatic oxidative stress and aminotransferases and initiated apoptosis, which may be related to the activated MAPK pathways in postweaning piglets. PMID:27807471

  14. Structural studies of Pseudomonas and Chromobacterium ω-aminotransferases provide insights into their differing substrate specificity.

    PubMed

    Sayer, Christopher; Isupov, Michail N; Westlake, Aaron; Littlechild, Jennifer A

    2013-04-01

    The crystal structures and inhibitor complexes of two industrially important ω-aminotransferase enzymes from Pseudomonas aeruginosa and Chromobacterium violaceum have been determined in order to understand the differences in their substrate specificity. The two enzymes share 30% sequence identity and use the same amino acceptor, pyruvate; however, the Pseudomonas enzyme shows activity towards the amino donor β-alanine, whilst the Chromobacterium enzyme does not. Both enzymes show activity towards S-α-methylbenzylamine (MBA), with the Chromobacterium enzyme having a broader substrate range. The crystal structure of the P. aeruginosa enzyme has been solved in the holo form and with the inhibitor gabaculine bound. The C. violaceum enzyme has been solved in the apo and holo forms and with gabaculine bound. The structures of the holo forms of both enzymes are quite similar. There is little conformational difference observed between the inhibitor complex and the holoenzyme for the P. aeruginosa aminotransferase. In comparison, the crystal structure of the C. violaceum gabaculine complex shows significant structural rearrangements from the structures of both the apo and holo forms of the enzyme. It appears that the different rigidity of the protein scaffold contributes to the substrate specificity observed for the two ω-aminotransferases.

  15. Structural studies of Pseudomonas and Chromobacterium ω-aminotransferases provide insights into their differing substrate specificity

    PubMed Central

    Sayer, Christopher; Isupov, Michail N.; Westlake, Aaron; Littlechild, Jennifer A.

    2013-01-01

    The crystal structures and inhibitor complexes of two industrially important ω-aminotransferase enzymes from Pseudomonas aeruginosa and Chromobacterium violaceum have been determined in order to understand the differences in their substrate specificity. The two enzymes share 30% sequence identity and use the same amino acceptor, pyruvate; however, the Pseudomonas enzyme shows activity towards the amino donor β-alanine, whilst the Chromobacterium enzyme does not. Both enzymes show activity towards S-α-methylbenzylamine (MBA), with the Chromobacterium enzyme having a broader substrate range. The crystal structure of the P. aeruginosa enzyme has been solved in the holo form and with the inhibitor gabaculine bound. The C. violaceum enzyme has been solved in the apo and holo forms and with gabaculine bound. The structures of the holo forms of both enzymes are quite similar. There is little conformational difference observed between the inhibitor complex and the holoenzyme for the P. aeruginosa aminotransferase. In comparison, the crystal structure of the C. violaceum gabaculine complex shows significant structural rearrangements from the structures of both the apo and holo forms of the enzyme. It appears that the different rigidity of the protein scaffold contributes to the substrate specificity observed for the two ω-­aminotransferases. PMID:23519665

  16. Identification and expression analyses of the alanine aminotransferase (AlaAT) gene family in poplar seedlings

    PubMed Central

    Xu, Zhiru; Ma, Jing; Qu, Chunpu; Hu, Yanbo; Hao, Bingqing; Sun, Yan; Liu, Zhongye; Yang, Han; Yang, Chengjun; Wang, Hongwei; Li, Ying; Liu, Guanjun

    2017-01-01

    Alanine aminotransferase (AlaAT, E.C.2.6.1.2) catalyzes the reversible conversion of pyruvate and glutamate to alanine and α-oxoglutarate. The AlaAT gene family has been well studied in some herbaceous plants, but has not been well characterized in woody plants. In this study, we identified four alanine aminotransferase homologues in Populus trichocarpa, which could be classified into two subgroups, A and B. AlaAT3 and AlaAT4 in subgroup A encode AlaAT, while AlaAT1 and AlaAT2 in subgroup B encode glutamate:glyoxylate aminotransferase (GGAT), which catalyzes the reaction of glutamate and glyoxylate to α-oxoglutarate and glycine. Four AlaAT genes were cloned from P. simonii × P. nigra. PnAlaAT1 and PnAlaAT2 were expressed predominantly in leaves and induced by exogenous nitrogen and exhibited a diurnal fluctuation in leaves, but was inhibited in roots. PnAlaAT3 and PnAlaAT4 were mainly expressed in roots, stems and leaves, and was induced by exogenous nitrogen. The expression of PnAlaAT3 gene could be regulated by glutamine or its related metabolites in roots. Our results suggest that PnAlaAT3 gene may play an important role in nitrogen metabolism and is regulated by glutamine or its related metabolites in the roots of P. simonii × P. nigra. PMID:28378825

  17. The aspartate aminotransferase-to-alanine aminotransferase ratio predicts all-cause and cardiovascular mortality in patients with type 2 diabetes

    PubMed Central

    Zoppini, Giacomo; Cacciatori, Vittorio; Negri, Carlo; Stoico, Vincenzo; Lippi, Giuseppe; Targher, Giovanni; Bonora, Enzo

    2016-01-01

    Abstract An increased aspartate aminotransferase-to-alanine aminotransferase ratio (AAR) has been widely used as a marker of advanced hepatic fibrosis. Increased AAR was also shown to be significantly associated with the risk of developing cardiovascular (CV) disease. The aim of this study was to assess the relationship between the AAR and mortality risk in a well-characterized cohort of patients with type 2 diabetes. A cohort of 2529 type 2 diabetic outpatients was followed-up for 6 years to collect cause-specific mortality. Cox regression analyses were modeled to estimate the independent association between AAR and the risk of all-cause and CV mortality. Over the 6-year follow-up period, 12.1% of patients died, 47.5% of whom from CV causes. An increased AAR, but not its individual components, was significantly associated with an increased risk of all-cause (adjusted-hazard risk 1.83, confidence interval [CI] 95% 1.14–2.93, P = 0.012) and CV (adjusted-hazard risk 2.60, CI 95% 1.38–4.90, P < 0.003) mortality after adjustment for multiple clinical risk factors and potential confounding variables. The AAR was independently associated with an increased risk of both all-cause and CV mortality in patients with type 2 diabetes. These findings suggest that an increased AAR may reflect more systemic derangements that are not simply limited to liver damage. Further studies are needed to elucidate the pathophysiological implications of an increased AAR. PMID:27787357

  18. Alanine Aminotransferase Elevation in Obese Infants and Children: A Marker of Early Onset Non Alcoholic Fatty Liver Disease

    PubMed Central

    Engelmann, Guido; Hoffmann, Georg Friedrich; Grulich-Henn, Juergen; Teufel, Ulrike

    2014-01-01

    Background: Elevated aminotransferases serve as surrogate markers of non-alcoholic fatty liver disease, a feature commonly associated with the metabolic syndrome. Studies on the prevalence of fatty liver disease in obese children comprise small patient samples or focus on those patients with liver enzyme elevation. Objectives: We have prospectively analyzed liver enzymes in all overweight and obese children coming to our tertiary care centre. Patients and Methods: In a prospective study 224 healthy, overweight or obese children aged 1 - 12 years were examined. Body Mass Index-Standard Deviation Score, alanine aminotransferase, aspartate aminotransferase and gamma-glutamyl-transpeptidase were measured. Results: Elevated alanine aminotransferase was observed in 29% of children. 26 % of obese and 30 % of overweight children had liver enzyme elevations. Obese children had significantly higher alanine aminotransferase levels than overweight children (0.9 vs. 0.7 times the Upper Limit of Normal; P = 0.04). Conclusions: Elevation of liver enzymes appears in 29 % obese children in a tertiary care centre. Absolute alanine aminotransferase levels are significantly higher in obese than in overweight children. Even obese children with normal liver enzymes show signs of fatty liver disease as demonstrated by liver enzymes at the upper limit of normal. PMID:24748893

  19. Thermal stability, pH dependence and inhibition of four murine kynurenine aminotransferases

    PubMed Central

    2010-01-01

    Background Kynurenine aminotransferase (KAT) catalyzes the transamination of kynunrenine to kynurenic acid (KYNA). KYNA is a neuroactive compound and functions as an antagonist of alpha7-nicotinic acetylcholine receptors and is the only known endogenous antagonist of N-methyl-D-aspartate receptors. Four KAT enzymes, KAT I/glutamine transaminase K/cysteine conjugate beta-lyase 1, KAT II/aminoadipate aminotransferase, KAT III/cysteine conjugate beta-lyase 2, and KAT IV/glutamic-oxaloacetic transaminase 2/mitochondrial aspartate aminotransferase, have been reported in mammalian brains. Because of the substrate overlap of the four KAT enzymes, it is difficult to assay the specific activity of each KAT in animal brains. Results This study concerns the functional expression and comparative characterization of KAT I, II, III, and IV from mice. At the applied test conditions, equimolar tryptophan with kynurenine significantly inhibited only mouse KAT I and IV, equimolar methionine inhibited only mouse KAT III and equimolar aspartate inhibited only mouse KAT IV. The activity of mouse KAT II was not significantly inhibited by any proteinogenic amino acids at equimolar concentrations. pH optima, temperature preferences of four KATs were also tested in this study. Midpoint temperatures of the protein melting, half life values at 65°C, and pKa values of mouse KAT I, II, III, and IV were 69.8, 65.9, 64.8 and 66.5°C; 69.7, 27.4, 3.9 and 6.5 min; pH 7.6, 5.7, 8.7 and 6.9, respectively. Conclusion The characteristics reported here could be used to develop specific assay methods for each of the four murine KATs. These specific assays could be used to identify which KAT is affected in mouse models for research and to develop small molecule drugs for prevention and treatment of KAT-involved human diseases. PMID:20482848

  20. Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia

    PubMed Central

    Abendroth, Jan; Choi, Ryan; Wall, Abigail; Clifton, Matthew C.; Lukacs, Christine M.; Staker, Bart L.; Van Voorhis, Wesley; Myler, Peter; Lorimer, Don D.; Edwards, Thomas E.

    2015-01-01

    The structures of three aspartate aminotransferases (AATs) from eukaryotic pathogens were solved within the Seattle Structural Genomics Center for Infectious Disease (SSGCID). Both the open and closed conformations of AAT were observed. Pyridoxal phosphate was bound to the active site via a Schiff base to a conserved lysine. An active-site mutant showed that Trypanosoma brucei AAT still binds pyridoxal phosphate even in the absence of the tethering lysine. The structures highlight the challenges for the structure-based design of inhibitors targeting the active site, while showing options for inhibitor design targeting the N-terminal arm. PMID:25945710

  1. Structures of aspartate aminotransferases from Trypanosoma brucei, Leishmania major and Giardia lamblia.

    PubMed

    Abendroth, Jan; Choi, Ryan; Wall, Abigail; Clifton, Matthew C; Lukacs, Christine M; Staker, Bart L; Van Voorhis, Wesley; Myler, Peter; Lorimer, Don D; Edwards, Thomas E

    2015-05-01

    The structures of three aspartate aminotransferases (AATs) from eukaryotic pathogens were solved within the Seattle Structural Genomics Center for Infectious Disease (SSGCID). Both the open and closed conformations of AAT were observed. Pyridoxal phosphate was bound to the active site via a Schiff base to a conserved lysine. An active-site mutant showed that Trypanosoma brucei AAT still binds pyridoxal phosphate even in the absence of the tethering lysine. The structures highlight the challenges for the structure-based design of inhibitors targeting the active site, while showing options for inhibitor design targeting the N-terminal arm.

  2. Inhibition study of alanine aminotransferase enzyme using sequential online capillary electrophoresis analysis.

    PubMed

    Liu, Lina; Chen, Yuanfang; Yang, Li

    2014-12-15

    We report the study of several inhibitors on alanine aminotransferase (ALT) enzyme using sequential online capillary electrophoresis (CE) assay. Using metal ions (Na(+) and Mg(2+)) as example inhibitors, we show that evolution of the ALT inhibition reaction can be achieved by automatically and simultaneously monitoring the substrate consumption and product formation as a function of reaction time. The inhibition mechanism and kinetic constants of ALT inhibition with succinic acid and two traditional Chinese medicines were derived from the sequential online CE assay. Our study could provide valuable information about the inhibition reactions of ALT enzyme.

  3. The anomalous kinetics of coupled aspartate aminotransferase and malate dehydrogenase. Evidence for compartmentation of oxaloacetate.

    PubMed Central

    Bryce, C F; Williams, D C; John, R A; Fasella, P

    1976-01-01

    Cytoplasmic aspartate aminotransferase and malate dehydrogenase were purified from pig heart. Kinetic parameters were determined for the separate reaction catalysed by each enzyme and used to predict the course of the coupled reaction: (see article). Although a lag phase should have been easily seen, none was detected. The same coupled reaction was also carried out by using radioactive aspartate in the presence of unlabelled oxaloacetate. The reaction was quenched with HClO4 after 70 ms and the specific radioactivity of the malate produced in this system was found to be essentially the same as that of the original aspartate. These results show that oxaloacetate produced by the aspartate aminotransferase is converted into malate by malate dehydrogenase before it equilibrates with the pool of unlabelled oxaloacetate and are consistent with a proposal that the enzymes are associated in a complex. However, no physical evidence of the existence of a complex could be found. An alternative means of compartmentation of the intermediate as an unstable isomer is considered. Images Fig. 2. Fig. 3. PMID:942372

  4. The past and present of serum aminotransferases and the future of liver injury biomarkers

    PubMed Central

    McGill, Mitchell R.

    2016-01-01

    Laboratory testing is important in the diagnosis and monitoring of liver injury and disease. Current liver tests include plasma markers of injury (e.g. aminotransferases, γ-glutamyl transferase, and alkaline phosphatase), markers of function (e.g. prothrombin time, bilirubin), viral hepatitis serologies, and markers of proliferation (e.g. α-fetoprotein). Among the injury markers, the alanine and aspartate aminotransferases (ALT and AST, respectively) are the most commonly used. However, interpretation of ALT and AST plasma levels can be complicated. Furthermore, both have poor prognostic utility in acute liver injury and liver failure. New biomarkers of liver injury are rapidly being developed, and the US Food and Drug Administration the European Medicines Agency have recently expressed support for use of some of these biomarkers in drug trials. The purpose of this paper is to review the history of liver biomarkers, to summarize mechanisms and interpretation of ALT and AST elevation in plasma in liver injury (particularly acute liver injury), and to discuss emerging liver injury biomarkers that may complement or even replace ALT and AST in the future. PMID:28337112

  5. The amino acid sequence of the aspartate aminotransferase from baker's yeast (Saccharomyces cerevisiae).

    PubMed Central

    Cronin, V B; Maras, B; Barra, D; Doonan, S

    1991-01-01

    1. The single (cytosolic) aspartate aminotransferase was purified in high yield from baker's yeast (Saccharomyces cerevisiae). 2. Amino-acid-sequence analysis was carried out by digestion of the protein with trypsin and with CNBr; some of the peptides produced were further subdigested with Staphylococcus aureus V8 proteinase or with pepsin. Peptides were sequenced by the dansyl-Edman method and/or by automated gas-phase methods. The amino acid sequence obtained was complete except for a probable gap of two residues as indicated by comparison with the structures of counterpart proteins in other species. 3. The N-terminus of the enzyme is blocked. Fast-atom-bombardment m.s. was used to identify the blocking group as an acetyl one. 4. Alignment of the sequence of the enzyme with those of vertebrate cytosolic and mitochondrial aspartate aminotransferases and with the enzyme from Escherichia coli showed that about 25% of residues are conserved between these distantly related forms. 5. Experimental details and confirmatory data for the results presented here are given in a Supplementary Publication (SUP 50164, 25 pages) that has been deposited at the British Library Document Supply Centre, Boston Spa. Wetherby, West Yorkshire LS23 7 BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1991) 273, 5. PMID:1859361

  6. Preoperative Aspartate Aminotransferase to White Blood Cell Count Ratio Predicting Postoperative Outcomes of Hepatocellular Carcinoma.

    PubMed

    Liao, Weijia; Wang, Yongqin; Liao, Yan; He, Songqing; Jin, Junfei

    2016-04-01

    Effective biomarkers for predicting prognosis of hepatocellular carcinoma (HCC) patients after hepatectomy is urgently needed. The purpose of this study is to evaluate the value of the preoperative peripheral aspartate aminotransferase to white blood cell count ratio (AWR) for the prognostication of patients with HCC.Clinical data of 396 HCC patients who underwent radical hepatectomy were retrospectively analyzed. The patients were divided into the low-AWR group (AWR ≤5.2) and the high-AWR group (AWR >5.2); univariate analysis, Kaplan-Meier method analysis, and the multivariate analysis by Cox regression were conducted, respectively.The results showed that AWR was associated with alpha-fetoprotein (AFP), tumor size, Barcelona clinic liver cancer (BCLC) stage, portal vein tumor thrombus (PVTT), and alanine aminotransferase (ALT) in HCC. AWR > 5.2, AFP > 100 ng/mL, size of tumor >6 cm, number of multiple tumors, B-C of BCLC stage, PVTT, and distant metastasis were predictors of poorer disease-free survival (DFS) and overall survival (OS). Except for recurrence, which was an independent predictor for OS only, AWR >5.2, size of tumor >6 cm, and PVTT were independent predictors of both DFS and OS.We concluded that preoperative AWR > 5.2 was an adverse predictor of DFS and OS in HCC after hepatectomy, AWR might be a novel prognostic biomarker in HCC after curative resection.

  7. An aminotransferase from Lactococcus lactis initiates conversion of amino acids to cheese flavor compounds.

    PubMed Central

    Yvon, M; Thirouin, S; Rijnen, L; Fromentier, D; Gripon, J C

    1997-01-01

    The enzymatic degradation of amino acids in cheese is believed to generate aroma compounds and therefore to be involved in the complex process of cheese flavor development. In lactococci, transamination is the first step in the degradation of aromatic and branched-chain amino acids which are precursors of aroma compounds. Here, the major aromatic amino acid aminotransferase of a Lactococcus lactis subsp. cremoris strain was purified and characterized. The enzyme transaminates the aromatic amino acids, leucine, and methionine. It uses the ketoacids corresponding to these amino acids and alpha-ketoglutarate as amino group acceptors. In contrast to most bacterial aromatic aminotransferases, it does not act on aspartate and does not use oxaloacetate as second substrate. It is essential for the transformation of aromatic amino acids to flavor compounds. It is a pyridoxal 5'-phosphate-dependent enzyme and is composed of two identical subunits of 43.5 kDa. The activity of the enzyme is optimal between pH 6.5 and 8 and between 35 and 45 degrees C, but it is still active under cheese-ripening conditions. PMID:9023921

  8. Prodynorphine opioid peptides and aspartate aminotransferase studied in spinal cord and sensory neurons

    SciTech Connect

    Sweetnam, P.M.

    1985-01-01

    An objective of this research was to obtain evidence for the synthesis and release of newly discovered opioid peptides, such as dynorphin, in spinal cord and sensory neurons. Several specific antisera were used to visualize dynorphin and related peptides in spinal cord and dorsal root ganglion neurons in dissociated cell culture. Antisera specific for the midportion of the dynorphin molecule revealed a subpopulation of spinal cord neurons with dense immunoreactive dynorphin in cell perikarya, but none in their associated neurites. Antisera specific for either the amino or carboxy terminal sequences of the molecule produced intense immunoreactivity in both cell perikarya and neurites of spinal neurons. These data suggest the cleavage products of dynorphin and not the complete molecule are possible neurotransmitters in the spinal cord. Additional evidence in support of this hypothesis was derived from radioimmunoassays of these cells and their culture medium following depolarization induced by elevated extracellular potassium. Antisera against aspartate aminotransferase revealed no differentially elevated immunoreactive aspartate aminotransferase in tissue sections of spinal cord or dorsal root ganglia.

  9. Pseudolinkage of the duplicate loci for supernatant aspartate aminotransferase in brook trout, Salvelinus fontinalis.

    PubMed

    Wright, J E; May, B; Stoneking, M; Lee, G M

    1980-01-01

    Electrophoretic variation involving three alleles is described for the duplicated loci for supernatant aspartate aminotransferase (AAT-1,2), from muscle extracts of brook trout. Both loci exhibit largely disomic inheritance. Exceptional progeny types are proposed to be the result of a form of tetrasomic inheritance. Nonrandom segregation was found among the progeny of males doubly heterozygous for AAT markers; where so-called linkage phase was known, this nonrandom assortment was shown to be pseudolinkage (78.9 percent recombination). Analyses of joint segregation of triply heterozygous males for the AAT-(1,2) loci and for the single alpha glycerophosphate dehydrogenase locus (AGP-1) revealed true linkage of AGP-1 with one AAT locus (mean r = 11 percent), but pseudolinkage with the other AAT locus (r = 74 percent). Intraindividual variation for homoeologous multivalent pairing of two acrocentric with two metacentric chromosomes in males, but with bivalent pairing in females, is proposed to account for pseudolinkage and for the tetrasomically inherited types.

  10. Hepatitis A virus genotype IA-infected patient with marked elevation of aspartate aminotransferase levels.

    PubMed

    Miura, Yoshifumi; Kanda, Tatsuo; Yasui, Shin; Takahashi, Koji; Haga, Yuki; Sasaki, Reina; Nakamura, Masato; Wu, Shuang; Nakamoto, Shingo; Arai, Makoto; Nishizawa, Tsutomu; Okamoto, Hiroaki; Yokosuka, Osamu

    2017-02-01

    We describe a case of acute liver failure (ALF) without hepatic encephalopathy with marked elevation of aminotransferase due to hepatitis A, according to the revised Japanese criteria of ALF. This liver biopsy of the patient showed compatible to acute viral hepatitis and she immediately recovered without intensive care. She had no comorbid disorders. Of interest, phylogenetic tree analysis using almost complete genomes of hepatitis A virus (HAV) demonstrated that the HAV isolate from her belonged to the HAV subgenotype IA strain and was similar to the HAJFF-Kan12 strain (99% nucleotide identity) or FH1 strain (98% nucleotide identity), which is associated with severe or fulminant hepatitis A. Careful interpretation of the association between HAV genome variations and severity of hepatitis A is needed and the mechanism of the severe hepatitis should be explored.

  11. Ornithine Aminotransferase, an Important Glutamate-Metabolizing Enzyme at the Crossroads of Multiple Metabolic Pathways

    PubMed Central

    Ginguay, Antonin; Cynober, Luc; Curis, Emmanuel; Nicolis, Ioannis

    2017-01-01

    Ornithine δ-aminotransferase (OAT, E.C. 2.6.1.13) catalyzes the transfer of the δ-amino group from ornithine (Orn) to α-ketoglutarate (aKG), yielding glutamate-5-semialdehyde and glutamate (Glu), and vice versa. In mammals, OAT is a mitochondrial enzyme, mainly located in the liver, intestine, brain, and kidney. In general, OAT serves to form glutamate from ornithine, with the notable exception of the intestine, where citrulline (Cit) or arginine (Arg) are end products. Its main function is to control the production of signaling molecules and mediators, such as Glu itself, Cit, GABA, and aliphatic polyamines. It is also involved in proline (Pro) synthesis. Deficiency in OAT causes gyrate atrophy, a rare but serious inherited disease, a further measure of the importance of this enzyme. PMID:28272331

  12. Effective disposal of nitrogen waste in blood-fed Aedes aegypti mosquitoes requires alanine aminotransferase

    PubMed Central

    Mazzalupo, Stacy; Isoe, Jun; Belloni, Virginia; Scaraffia, Patricia Y.

    2016-01-01

    To better understand the mechanisms responsible for the success of female mosquitoes in their disposal of excess nitrogen, we investigated the role of alanine aminotransferase (ALAT) in blood-fed Aedes aegypti. Transcript and protein levels from the 2 ALAT genes were analyzed in sucrose- and blood-fed A. aegypti tissues. ALAT1 and ALAT2 exhibit distinct expression patterns in tissues during the first gonotrophic cycle. Injection of female mosquitoes with either double-stranded RNA (dsRNA)-ALAT1 or dsRNA ALAT2 significantly decreased mRNA and protein levels of ALAT1 or ALAT2 in fat body, thorax, and Malpighian tubules compared with dsRNA firefly luciferase-injected control mosquitoes. The silencing of either A. aegypti ALAT1 or ALAT2 caused unexpected phenotypes such as a delay in blood digestion, a massive accumulation of uric acid in the midgut posterior region, and a significant decrease of nitrogen waste excretion during the first 48 h after blood feeding. Concurrently, the expression of genes encoding xanthine dehydrogenase and ammonia transporter (Rhesus 50 glycoprotein) were significantly increased in tissues of both ALAT1- and ALAT2-deficient females. Moreover, perturbation of ALAT1 and ALAT2 in the female mosquitoes delayed oviposition and reduced egg production. These novel findings underscore the efficient mechanisms that blood-fed mosquitoes use to avoid ammonia toxicity and free radical damage.—Mazzalupo, S., Isoe, J., Belloni, V., Scaraffia, P. Y. Effective disposal of nitrogen waste in blood-fed Aedes aegypti mosquitoes requires alanine aminotransferase. PMID:26310269

  13. DL-canaline and 5-fluoromethylornithine. Comparison of two inactivators of ornithine aminotransferase.

    PubMed Central

    Bolkenius, F N; Knödgen, B; Seiler, N

    1990-01-01

    5-Fluoromethylornithine (5FMOrn) is an enzyme-activated irreversible inhibitor or ornithine aminotransferase (L-ornithine:2-oxo-acid 5-aminotransferase, OAT). For purified rat liver OAT, Ki(app.) was found to be 30 microM. and tau 1/2 = 4 min. Of the four stereomers of 5FMOrn only one reacts with OAT. The formation of a chromophore with an absorption maximum at 458 nm after inactivation of OAT by 5FMOrn suggests the formation of an enamine intermediate, which is slowly hydrolysed to release an unsaturated ketone. L-Canaline [(S)-2-amino-4-amino-oxybutyric acid] is a well-known irreversible inhibitor of OAT. Not only the natural L-enantiomer but also the D-enantiomer reacts by oxime formation with pyridoxal 5'-phosphate in the active site of the enzyme, although considerably more slowly. This demonstrates that the stereochemistry at C-2 of ornithine is not absolutely stringent. In vitro, canaline reacted faster than 5FMOrn with OAT. In vivo, however, only incomplete OAT inhibition was observed with canaline. Whereas intraperitoneal administration of 10 mg of 5FMOrn/kg body wt. to mice was sufficient to inactivate OAT in brain and liver by 90% for 24 h, 500 mg of DL-canaline/kg body wt. only produced a transient inhibition of 65-70%. The accumulation of ornithine in these tissues was considerably slower and the maximum concentrations lower than were achieved with 5FMOrn. It appears that DL-canaline, in contrast with 5FMOrn, is not useful as a tool in studies of biological consequences of OAT inhibition. PMID:2363680

  14. Irritable Bowel Syndrome May Be Associated with Elevated Alanine Aminotransferase and Metabolic Syndrome

    PubMed Central

    Lee, Seung-Hwa; Kim, Kwang-Min; Joo, Nam-Seok

    2016-01-01

    Purpose Recent studies have revealed close relationships between hepatic injury, metabolic pathways, and gut microbiota. The microorganisms in the intestine also cause irritable bowel syndrome (IBS). The aim of this study was to examine whether IBS was associated with elevated hepatic enzyme [alanine aminotransferase (ALT) and aspartate aminotransferase (AST)], gamma-glutamyl transferase (γ-GT) levels, and metabolic syndrome (MS). Materials and Methods This was a retrospective, cross-sectional, case-control study. The case and control groups comprised subjects who visited our health promotion center for general check-ups from June 2010 to December 2010. Of the 1127 initially screened subjects, 83 had IBS according to the Rome III criteria. The control group consisted of 260 age- and sex-matched subjects without IBS who visited our health promotion center during the same period. Results Compared to control subjects, patients with IBS showed significantly higher values of anthropometric parameters (body mass index, waist circumference), liver enzymes, γ-GT, and lipid levels. The prevalences of elevated ALT (16.9% vs. 7.7%; p=0.015) and γ-GT (24.1% vs. 11.5%; p=0.037) levels were significantly higher in patients with IBS than in control subjects. A statistically significant difference was observed in the prevalence of MS between controls and IBS patients (12.7% vs. 32.5%; p<0.001). The relationships between elevated ALT levels, MS, and IBS remained statistically significant after controlling for potential confounding factors. Conclusion On the basis of our study results, IBS may be an important condition in certain patients with elevated ALT levels and MS. PMID:26632395

  15. Histidine degradation via an aminotransferase increases the nutritional flexibility of Candida glabrata.

    PubMed

    Brunke, Sascha; Seider, Katja; Richter, Martin Ernst; Bremer-Streck, Sibylle; Ramachandra, Shruthi; Kiehntopf, Michael; Brock, Matthias; Hube, Bernhard

    2014-06-01

    The ability to acquire nutrients during infections is an important attribute in microbial pathogenesis. Amino acids are a valuable source of nitrogen if they can be degraded by the infecting organism. In this work, we analyzed histidine utilization in the fungal pathogen of humans Candida glabrata. Hemiascomycete fungi, like C. glabrata or Saccharomyces cerevisiae, possess no gene coding for a histidine ammonia-lyase, which catalyzes the first step of a major histidine degradation pathway in most other organisms. We show that C. glabrata instead initializes histidine degradation via the aromatic amino acid aminotransferase Aro8. Although ARO8 is also present in S. cerevisiae and is induced by extracellular histidine, the yeast cannot use histidine as its sole nitrogen source, possibly due to growth inhibition by a downstream degradation product. Furthermore, C. glabrata relies only on Aro8 for phenylalanine and tryptophan utilization, since ARO8, but not its homologue ARO9, was transcriptionally activated in the presence of these amino acids. Accordingly, an ARO9 deletion had no effect on growth with aromatic amino acids. In contrast, in S. cerevisiae, ARO9 is strongly induced by tryptophan and is known to support growth on aromatic amino acids. Differences in the genomic structure of the ARO9 gene between C. glabrata and S. cerevisiae indicate a possible disruption in the regulatory upstream region. Thus, we show that, in contrast to S. cerevisiae, C. glabrata has adapted to use histidine as a sole source of nitrogen and that the aromatic amino acid aminotransferase Aro8, but not Aro9, is the enzyme required for this process.

  16. Abdominal obesity validates the association between elevated alanine aminotransferase and newly diagnosed diabetes mellitus.

    PubMed

    Yueh, Chen-Yu; Yang, Yao-Hsu; Sung, Yi-Ting; Lee, Li-Wen

    2014-01-01

    To examine how elevated alanine aminotransferase (ALT) could be associated with newly diagnosed diabetes mellitus. We conducted a cross-sectional analysis on a mass health examination. The odds ratios (ORs) for diabetes mellitus and newly diagnosed diabetes mellitus were compared between people with and without abdominal obesity, together with and without elevated ALT levels. 5499 people were included in this study. Two hundred fifty two (4.6%) fulfilled the diagnosis of diabetes mellitus with 178 (3.2%) undiagnosed before. Metabolic syndrome was vigorously associated with diabetes mellitus and newly diagnosed diabetes mellitus (12.4% vs. 1.4% and 9.0% vs. 0.9%), but elevated ALT alone was not. However, coexisting with obesity, elevated ALTs were robustly associated with diabetes mellitus and newly diagnosed diabetes mellitus. For the incidence of newly diagnosed diabetes mellitus, in comparison to non-obese people with normal ALT (1.7%, OR = 1), obese people especially with elevated ALT levels had significantly higher ORs (obese with ALT ≤ 40 U/L: 4.7%, OR 1.73, 95% CI 1.08-2.77, P 0.023; ALT 41-80 U/L: 6.8%, OR 2.06, 95% CI 1.20-3.55, P 0.009; ALT 81-120 U/L: 8.8%, OR 3.07, 95% CI 1.38-6.84, P 0.006; ALT > 120 U/L: 18.2%, OR 7.44, 95% CI 3.04-18.18, P < 0.001). Abdominal obesity validates the association between elevated alanine aminotransferase and diabetes mellitus and newly diagnosed diabetes mellitus. People with abdominal obesity, especially with coexisting elevated ALT levels should be screened for undiagnosed diabetes mellitus.

  17. Phenylalanine-pyruvate aminotransferase activity in chicks subjected to phenylalanine imbalance or phenylalanine toxicity.

    PubMed

    Lu, J; Austic, R E

    2009-11-01

    Two experiments were done to determine the influence of Phe imbalance and excess on Phe-pyruvate aminotransferase (PAT) activity in the chick. Five replicates of 3 chicks (experiment 1) or 2 chicks (experiment 2) of a commercial brown egg layer strain were fed a semipurified diet for 1 wk and then received experimental diets for 10 d. Three diets were used in experiment 1: the basal diet contained 0.46% Phe; the imbalance diet was similar to the basal diet except that it contained a 10% mixture of indispensable amino acids lacking Phe (IAA - Phe) to create a Phe imbalance; the imbalance corrected diet was similar to the imbalance diet except that it was supplemented with 1.12% Phe to correct the imbalance. A 3 x 2 factorial arrangement of treatments in experiment 2 provided 3 dietary levels (0.46, 1.58, and 2.46%) of Phe and either no supplement or 10% supplement of IAA - Phe. Nonfasted chicks were killed and livers were sampled in experiment 1, and livers, kidneys, brains, and pectoralis major muscles were sampled in experiment 2. In experiment 1, liver PAT activity per gram of liver was 80 and 55% higher (P < 0.01) in chicks fed the imbalance and imbalance corrected diets than in chicks fed the basal diet. In experiment 2, the livers and kidneys, but not brains and muscles, of chicks that received the 10% supplement of IAA - Phe had higher activities of PAT per gram of tissue per minute and per milligram of tissue protein extract per minute than chicks that did not receive IAA - Phe (P < 0.001). No effect of dietary Phe on PAT activity was detected (P > 0.05). Phenylalanine-pyruvate aminotransferase activity appears to be regulated in response to dietary content of indispensable amino acids but not by the dietary level of Phe.

  18. The effect of portacaval anastomosis on the expression of glutamine synthetase and ornithine aminotransferase in perivenous hepatocytes.

    PubMed

    da Silva, Robin; Levillain, Oliver; Brosnan, John T; Araneda, Silvia; Brosnan, Margaret E

    2013-05-01

    There is functional zonation of metabolism across the liver acinus, with glutamine synthetase restricted to a narrow band of cells around the terminal hepatic venules. Portacaval anastomosis, where there is a major rerouting of portal blood flow from the portal vein directly to the vena cava bypassing the liver, has been reported to result in a marked decrease in the activity of glutamine synthetase. It is not known whether this represents a loss of perivenous hepatocytes or whether there is a specific loss of glutamine synthetase. To answer this question, we have determined the activity of glutamine synthetase and another enzyme from the perivenous compartment, ornithine aminotransferase, as well as the immunochemical localization of both glutamine synthetase and ornithine aminotransferase in rats with a portacaval shunt. The portacaval shunt caused a marked decrease in glutamine synthetase activity and an increase in ornithine aminotransferase activity. Immunohistochemical analysis showed that the glutamine synthetase and ornithine aminotransferase proteins maintained their location in the perivenous cells. These results indicate that there is no generalized loss of perivenous hepatocytes, but rather, there is a significant alteration in the expression of these proteins and hence metabolism in this cell population.

  19. Paralogous ALT1 and ALT2 Retention and Diversification Have Generated Catalytically Active and Inactive Aminotransferases in Saccharomyces cerevisiae

    PubMed Central

    Peñalosa-Ruiz, Georgina; Aranda, Cristina; Ongay-Larios, Laura; Colon, Maritrini; Quezada, Hector; Gonzalez, Alicia

    2012-01-01

    Background Gene duplication and the subsequent divergence of paralogous pairs play a central role in the evolution of novel gene functions. S. cerevisiae possesses two paralogous genes (ALT1/ALT2) which presumably encode alanine aminotransferases. It has been previously shown that Alt1 encodes an alanine aminotransferase, involved in alanine metabolism; however the physiological role of Alt2 is not known. Here we investigate whether ALT2 encodes an active alanine aminotransferase. Principal Findings Our results show that although ALT1 and ALT2 encode 65% identical proteins, only Alt1 displays alanine aminotransferase activity; in contrast ALT2 encodes a catalytically inert protein. ALT1 and ALT2 expression is modulated by Nrg1 and by the intracellular alanine pool. ALT1 is alanine-induced showing a regulatory profile of a gene encoding an enzyme involved in amino acid catabolism, in agreement with the fact that Alt1 is the sole pathway for alanine catabolism present in S. cerevisiae. Conversely, ALT2 expression is alanine-repressed, indicating a role in alanine biosynthesis, although the encoded-protein has no alanine aminotransferase enzymatic activity. In the ancestral-like yeast L. kluyveri, the alanine aminotransferase activity was higher in the presence of alanine than in the presence of ammonium, suggesting that as for ALT1, LkALT1 expression could be alanine-induced. ALT2 retention poses the questions of whether the encoded protein plays a particular function, and if this function was present in the ancestral gene. It could be hypotesized that ALT2 diverged after duplication, through neo-functionalization or that ALT2 function was present in the ancestral gene, with a yet undiscovered function. Conclusions ALT1 and ALT2 divergence has resulted in delegation of alanine aminotransferase activity to Alt1. These genes display opposed regulatory profiles: ALT1 is alanine-induced, while ALT2 is alanine repressed. Both genes are negatively regulated by the Nrg1

  20. Elevated Aspartate and Alanine Aminotransferase Levels and Natural Death among Patients with Methamphetamine Dependence

    PubMed Central

    Kuo, Chian-Jue; Tsai, Shang-Ying; Liao, Ya-Tang; Conwell, Yeates; Lee, Wen-Chung; Huang, Ming-Chyi; Lin, Shih-Ku; Chen, Chiao-Chicy; Chen, Wei J.

    2012-01-01

    Background Methamphetamine is one of the fastest growing illicit drugs worldwide, causing multiple organ damage and excessive natural deaths. The authors aimed to identify potential laboratory indices and clinical characteristics associated with natural death through a two-phase study. Methods Methamphetamine-dependent patients (n = 1,254) admitted to a psychiatric center in Taiwan between 1990 and 2007 were linked with a national mortality database for causes of death. Forty-eight subjects died of natural causes, and were defined as the case subjects. A time-efficient sex- and age-matched nested case-control study derived from the cohort was conducted first to explore the potential factors associated with natural death through a time-consuming standardized review of medical records. Then the identified potential factors were evaluated in the whole cohort to validate the findings. Results In phase I, several potential factors associated with natural death were identified, including aspartate aminotransferase (AST), alanine aminotransferase (ALT), comorbid alcohol use disorder, and the prescription of antipsychotic drugs. In phase II, these factors were confirmed in the whole cohort using survival analysis. For the characteristics at the latest hospital admission, Cox proportional hazards models showed that the adjusted hazard ratios for natural death were 6.75 (p<0.001) in the group with markedly elevated AST (>80 U/L) and 2.66 (p<0.05) in the group with mildly elevated AST (40–80 U/L), with reference to the control group (<40 U/L). As for ALT, the adjusted hazard ratios were 5.41 (p<0.001), and 1.44 (p>0.05). Comorbid alcohol use disorder was associated with an increased risk of natural death, whereas administration of antipsychotic drugs was not associated with lowered risk. Conclusions This study highlights the necessity of intensive follow-up for those with elevated AST and ALT levels and comorbid alcohol use disorder for preventing excessive natural

  1. Cloning and molecular characterization of tick kynurenine aminotransferase (HlKAT) from Haemaphysalis longicornis (Acari: Ixodidae).

    PubMed

    Battsetseg, Badgar; Boldbaatar, Damdinsuren; Battur, Banzragch; Xuan, Xuenan; Fujisaki, Kozo

    2009-09-01

    A complementary DNA coding a novel kynurenine aminotransferase (KAT) molecule from Haemaphysalis longicornis tick embryo was cloned and characterized. The transcription of the HlKAT occurs at all stages during tick development as well as in the midgut, salivary glands, ovary, and synganglion of adult ticks, and protein expression levels increased during the blood-feeding course. The HlKAT gene without signal peptide was successfully expressed as a glutathione S-transferase fusion protein in soluble form, which is capable of catalyzing the transamination of kynurenine and 3-hydroxykynurenine to kynurenic acid and xanthurenic acid, respectively. The purified recombinant HlKAT showed dose-dependent inhibition effect on the growth of equine babesial parasite, Babesia caballi, in in vitro culture. All results suggested that a specific HlKAT is present in tick and HlKAT may play an important physiological role in H. longicornis. This is the first report of a member enzyme of tryptophan pathway in Chelicerata.

  2. Structure of GroEL in Complex with an Early Folding Intermediate of Alanine Glyoxylate Aminotransferase*

    PubMed Central

    Albert, Armando; Yunta, Cristina; Arranz, Rocío; Peña, Álvaro; Salido, Eduardo; Valpuesta, José María; Martín-Benito, Jaime

    2010-01-01

    Primary hyperoxaluria type 1 is a rare autosomal recessive disease caused by mutations in the alanine glyoxylate aminotransferase gene (AGXT). We have previously shown that P11L and I340M polymorphisms together with I244T mutation (AGXT-LTM) represent a conformational disease that could be amenable to pharmacological intervention. Thus, the study of the folding mechanism of AGXT is crucial to understand the molecular basis of the disease. Here, we provide biochemical and structural data showing that AGXT-LTM is able to form non-native folding intermediates. The three-dimensional structure of a complex between the bacterial chaperonin GroEL and a folding intermediate of AGXT-LTM mutant has been solved by cryoelectron microscopy. The electron density map shows the protein substrate in a non-native extended conformation that crosses the GroEL central cavity. Addition of ATP to the complex induces conformational changes on the chaperonin and the internalization of the protein substrate into the folding cavity. The structure provides a three-dimensional picture of an in vivo early ATP-dependent step of the folding reaction cycle of the chaperonin and supports a GroEL functional model in which the chaperonin promotes folding of the AGXT-LTM mutant protein through forced unfolding mechanism. PMID:20056599

  3. Structural insight into the inhibition of human kynurenine aminotransferase I/glutamine transaminase K.

    PubMed

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A; Li, Jianyong

    2009-05-14

    Human kynurenine aminotransferase I (hKAT I) catalyzes the formation of kynurenic acid, a neuroactive compound. Here, we report three high-resolution crystal structures (1.50-1.55 A) of hKAT I that are in complex with glycerol and each of two inhibitors of hKAT I: indole-3-acetic acid (IAC) and Tris. Because Tris is able to occupy the substrate binding position, we speculate that this may be the basis for hKAT I inhibition. Furthermore, the hKAT/IAC complex structure reveals that the binding moieties of the inhibitor are its indole ring and a carboxyl group. Six chemicals with both binding moieties were tested for their ability to inhibit hKAT I activity; 3-indolepropionic acid and DL-indole-3-lactic acid demonstrated the highest level of inhibition, and as they cannot be considered as substrates of the enzyme, these two inhibitors are promising candidates for future study. Perhaps even more significantly, we report the discovery of two different ligands located simultaneously in the hKAT I active center for the first time.

  4. Regulation of 2-oxoglutarate metabolism in rat liver by NADP-isocitrate dehydrogenase and aspartate aminotransferase.

    PubMed

    Rakhmanova, T I; Popova, T N

    2006-02-01

    Kinetic and regulatory properties of NADP-isocitrate dehydrogenase (NADP-IDH) and aspartate aminotransferase (AsAT) responsible for 2-oxoglutarate metabolism in the cytoplasm and mitochondria of rat liver were studied. Based on the subcellular location of these enzymes and their kinetic parameters (Km, Ksi) obtained with highly purified enzyme preparations, it is suggested that synthesis of 2-oxoglutarate should be mainly determined by cytoplasmic NADP-IDH (86% of the total activity in the cell), whereas its utilization should depend on cytoplasmic AsAT (78% of the total activity). AsAT from the rat liver was specified by substrate inhibition and also by changes in the enzyme affinity for the substrates under the influence of some intermediates of the tricarboxylic acid cycle: isocitrate, succinate, fumarate, and citrate. Key intermediates of nitrogen metabolism (glutamate, glutamine, and aspartate) are involved in the regulation of NADP-IDH and AsAT. These enzymes are regulated oppositely, and the catalytic activity of one enzyme can be stimulated concurrently with a decrease in the activity of the other. Obviously, carbon and nitrogen metabolism in the rat liver can be controlled through redistribution of 2-oxoglutarate between different metabolic processes via regulatory mechanisms influencing differently located forms of NADP-IDH and AsAT.

  5. Structural Insight into the Inhibition of Human Kynurenine Aminotransferase I/Glutamine transaminase K∥

    PubMed Central

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A.; Li, Jianyong

    2010-01-01

    Human kynurenine aminotransferase I (hKAT I) catalyzes the formation of kynurenic acid, a neuroactive compound. Here, we report three high-resolution crystal structures (1.50–1.55 Å) of hKAT I that are in complex with glycerol and each of two inhibitors of hKAT I: indole-3-acetic acid (IAC) and Tris. Because Tris is able to occupy the substrate binding position, we speculate that this may be the basis for hKAT I inhibition. Furthermore, the hKAT/IAC complex structure reveals that the binding moieties of the inhibitor are its indole ring and a carboxyl group. Six chemicals with both binding moieties were tested for their ability to inhibit hKAT I activity; 3-indolepropionic acid and DL-indole-3-lactic acid demonstrated the highest level of inhibition, and as they cannot be considered as substrates of the enzyme, these two inhibitors are promising candidates for future study. Perhaps even more significantly, we report the discovery of two different ligands located simultaneously in the hKAT I active center for the first time. PMID:19338303

  6. Structural Basis for the Stereochemical Control of Amine Installation in Nucleotide Sugar Aminotransferases.

    PubMed

    Wang, Fengbin; Singh, Shanteri; Xu, Weijun; Helmich, Kate E; Miller, Mitchell D; Cao, Hongnan; Bingman, Craig A; Thorson, Jon S; Phillips, George N

    2015-09-18

    Sugar aminotransferases (SATs) are an important class of tailoring enzymes that catalyze the 5'-pyridoxal phosphate (PLP)-dependent stereo- and regiospecific installation of an amino group from an amino acid donor (typically L-Glu or L-Gln) to a corresponding ketosugar nucleotide acceptor. Herein we report the strategic structural study of two homologous C4 SATs (Micromonospora echinospora CalS13 and Escherichia coli WecE) that utilize identical substrates but differ in their stereochemistry of aminotransfer. This study reveals for the first time a new mode of SAT sugar nucleotide binding and, in conjunction with previously reported SAT structural studies, provides the basis from which to propose a universal model for SAT stereo- and regiochemical control of amine installation. Specifically, the universal model put forth highlights catalytic divergence to derive solely from distinctions within nucleotide sugar orientation upon binding within a relatively fixed SAT active site where the available ligand bound structures of the three out of four representative C3 and C4 SAT examples provide a basis for the overall model. Importantly, this study presents a new predictive model to support SAT functional annotation, biochemical study and rational engineering.

  7. Molecular basis of ornithine aminotransferase deficiency in B-6-responsive and -nonresponsive forms of gyrate atrophy

    SciTech Connect

    Ramesh, V.; McClatchey, A.I.; Ramesh, N.; Benoit, L.A.; Berson, E.L.; Shih, V.E.; Gusella, J.F. )

    1988-06-01

    Gyrate atrophy (GA), a recessive eye disease involving progressive loss of vision due to chorioretinal degeneration, is associated with a deficiency of the mitochondrial enzyme ornithine aminotransferase with consequent hyperornithinemia. Genetic heterogeneity of GA has been suggested by the demonstration that administration of pyridoxine to increase the level of pyridoxal phosphate, a cofactor of OATase, reduces hyperornithinemia in a subset of patients. The authors have cloned and sequences cDNAs for OATase from two GA patients, one responsive and one nonresponsive to pyridoxine treatment. The respective cDNAs contained different single missense mutations, which were sufficient to eliminate OATase activity when each cDNA was tested in a eukaryotic expression system. However, like the enzyme in fibroblasts from the pyridoxine-responsive patient, OATase encoded by the corresponding cDNA from this individual showed a significant increase in activity when assayed in the presence of an increased pyridoxal phosphate concentration. These data firmly establish that both pyridoxine responsive and nonresponsive forms of GA result from mutations in the OATase structural gene. Moreover, they provide a molecular characterization of the primary lesion in a pyridoxine-responsive genetic disorder.

  8. CPP-115, a Potent γ-Aminobutyric Acid Aminotransferase Inactivator for the Treatment of Cocaine Addiction

    PubMed Central

    Pan, Yue; Gerasimov, Madina R.; Kvist, Trine; Wellendorph, Petrine; Madsen, Karsten K.; Pera, Elena; Lee, Hyunbeom; Schousboe, Arne; Chebib, Mary; Bräuner-Osborne, Hans; Craft, Cheryl M.; Brodie, Jonathan D.; Schiffer, Wynne K.; Dewey, Stephen L.; Miller, Steven R.; Silverman, Richard B.

    2011-01-01

    Vigabatrin, a GABA aminotransferase (GABA-AT) inactivator, is used to treat infantile spasms and refractory complex partial seizures and is in clinical trials to treat addiction. We evaluated a novel GABA-AT inactivator (CPP-115) and observed that it does not exhibit other GABAergic or off-target activities and is rapidly and completely orally absorbed and eliminated. Using in vivo microdialysis techniques in freely moving rats and micro-PET imaging techniques, CPP-115 produced similar inhibition of cocaine-induced increases in extracellular dopamine and in synaptic dopamine in the nucleus accumbens at 1/300–1/600th the dose of vigabatrin. It also blocks expression of cocaine-induced conditioned place preference at a dose 1/300th that of vigabatrin. Electroretinographic (ERG) responses in rats treated with CPP-115, at doses 20–40 times higher than those needed to treat addiction in rats, exhibited reductions in ERG responses, which were less than the reductions observed in rats treated with vigabatrin at the same dose needed to treat addiction in rats. In conclusion, CPP-115 can be administered at significantly lower doses than vigabatrin, which suggests a potential new treatment for addiction with a significantly reduced risk of visual field defects. PMID:22128851

  9. Effects of self-association of ornithine aminotransferase on its physicochemical characteristics

    SciTech Connect

    Boernke, W.E.; Stevens, F.J.; Peraino, C.

    1981-01-01

    Previous work in this laboratory has shown that the molecular weight of ornithine aminotransferase (OAT) is concentration dependent. In the present study this property of OAT was further characterized by using sedimentation equilibrium centrifugation to determine the molecular weight of OAT in a range of enzyme concentrations. It was shown that OAT aggregates in a two-stage process as its concentration increases. The first stage involves the association of enzymatically active monomers into trimers, with association of the trimmers into higher order aggregates occurring in the second stage. Decreasing the pH or raising the ionic strength enhanced aggregation, while raising the pH inhibits aggregation; however, the two-stage nature of the aggregation process was not affected by changes in pH and ionic strength. Kinetic analyses of purified enzyme showed that aggregatio results in an increase in the K/sub m/ for both substrates with the V/sub max/ remaining constant, indicating that aggregation of monomers sterically hinders substrate binding. Increased K/sub m/ values were also obtained for OAT sequestered in mitochondia from rats fed a high-protein diet to increase mitochondrial OAT levels. The higher K/sub m/ values suggest that the elevation of OAT in vivo is accompanied by aggregation of the enzyme within the mitochondrion. We propose that the aggregation-dependent increase of K/sub m/ in vivo has adaptive value in that it spares ornithine for use in the urea cycle.

  10. The unfolding pathway for Apo Escherichia coli aspartate aminotransferase is dependent on the choice of denaturant.

    PubMed

    Deu, Edgar; Kirsch, Jack F

    2007-05-15

    The guanidine hydrochloride (GdnHCl) mediated denaturation pathway for the apo form of homodimeric Escherichia coli aspartate aminotransferase (eAATase) (molecular mass = 43.5 kDa/monomer) includes a partially folded monomeric intermediate, M* [Herold, M., and Kirschner, K. (1990) Biochemistry 29, 1907-1913; Birolo, L., Dal Piaz, F., Pucci, P., and Marino, G. (2002) J. Biol. Chem. 277, 17428-17437]. The present investigation of the urea-mediated denaturation of eAATase finds no evidence for an M* species but uncovers a partially denatured dimeric form, D*, that is unpopulated in GdnHCl. Thus, the unfolding process is a function of the employed denaturant. D* retains less than 50% of the native secondary structure (circular dichroism), conserves significant quaternary and tertiary interactions, and unfolds cooperatively (mD*<==>U = 3.4 +/- 0.3 kcal mol-1 M-1). Therefore, the following equilibria obtain in the denaturation of apo-eAATase: D <==> 2M 2M* <==> 2U in GdnHCl and D <==> D* <==> 2U in urea (D = native dimer, M = folded monomer, and U = unfolded state). The free energy of unfolding of apo-eAATase (D <==> 2U) is 36 +/- 3 kcal mol-1, while that for the D* 2U transition is 24 +/- 2 kcal mol-1, both at 1 M standard state and pH 7.5.

  11. Aspartate aminotransferase activity in the pulp of teeth treated for 6 months with fixed orthodontic appliances

    PubMed Central

    Latkauskiene, Dalia; Racinskaite, Vilma; Skucaite, Neringa; Machiulskiene, Vita

    2015-01-01

    Objective To measure aspartate aminotransferase (AST) activity in the pulp of teeth treated with fixed appliances for 6 months, and compare it with AST activity measured in untreated teeth. Methods The study sample consisted of 16 healthy subjects (mean age 25.7 ± 4.3 years) who required the extraction of maxillary premolars for orthodontic reasons. Of these, 6 individuals had a total of 11 sound teeth extracted without any orthodontic treatment (the control group), and 10 individuals had a total of 20 sound teeth extracted after 6 months of orthodontic alignment (the experimental group). Dental pulp samples were extracted from all control and experimental teeth, and the AST activity exhibited by these samples was determined spectrophotometrically at 20℃. Results Mean AST values were 25.29 × 10-5 U/mg (standard deviation [SD] 9.95) in the control group and 27.54 × 10-5 U/mg (SD 31.81) in the experimental group. The difference between these means was not statistically significantly (p = 0.778), and the distribution of the AST values was also similar in both groups. Conclusions No statistically significant increase in AST activity in the pulp of mechanically loaded teeth was detected after 6 months of orthodontic alignment, as compared to that of teeth extracted from individuals who had not undergone orthodontic treatment. This suggests that time-related regenerative processes occur in the dental pulp. PMID:26445721

  12. Bicyclic γ-amino acids as inhibitors of γ-aminobutyrate aminotransferase.

    PubMed

    Pinto, Andrea; Tamborini, Lucia; Pennacchietti, Eugenia; Coluccia, Antonio; Silvestri, Romano; Cullia, Gregorio; De Micheli, Carlo; Conti, Paola; De Biase, Daniela

    2016-01-01

    The γ-aminobutyrate (GABA)-degradative enzyme GABA aminotransferase (GABA-AT) is regarded as an attractive target to control GABA levels in the central nervous system: this has important implications in the treatment of several neurological disorders and drug dependencies. We have investigated the ability of newly synthesized compounds to act as GABA-AT inhibitors. These compounds have a unique bicyclic structure: the carbocyclic ring bears the GABA skeleton, while the fused 3-Br-isoxazoline ring contains an electrophilic warhead susceptible of nucleophilic attack by an active site residue of the target enzyme. Out of the four compounds tested, only the one named (+)-3 was found to significantly inhibit mammalian GABA-AT in vitro. Docking studies, performed on the available structures of GABA-AT, support the experimental findings: out of the four tested compounds, only (+)-3 suitably orients the electrophilic 3-Br-isoxazoline warhead towards the active site nucleophilic residue Lys329, thereby explaining the irreversible inhibition of GABA-AT observed experimentally.

  13. A Micro-Platinum Wire Biosensor for Fast and Selective Detection of Alanine Aminotransferase

    PubMed Central

    Thuy, Tran Nguyen Thanh; Tseng, Tina T.-C.

    2016-01-01

    In this study, a miniaturized biosensor based on permselective polymer layers (overoxidized polypyrrole (Ppy) and Nafion®) modified and enzyme (glutamate oxidase (GlutOx)) immobilized micro-platinum wire electrode for the detection of alanine aminotransferase (ALT) was fabricated. The proposed ALT biosensor was measured electrochemically by constant potential amperometry at +0.7 V vs. Ag/AgCl. The ALT biosensor provides fast response time (~5 s) and superior selectivity towards ALT against both negatively and positively charged species (e.g., ascorbic acid (AA) and dopamine (DA), respectively). The detection range of the ALT biosensor is found to be 10–900 U/L which covers the range of normal ALT levels presented in the serum and the detection limit and sensitivity are found to be 8.48 U/L and 0.059 nA/(U/L·mm2) (N = 10), respectively. We also found that one-day storage of the ALT biosensor at −20 °C right after the sensor being fabricated can enhance the sensor sensitivity (1.74 times higher than that of the sensor stored at 4 °C). The ALT biosensor is stable after eight weeks of storage at −20 °C. The sensor was tested in spiked ALT samples (ALT activities: 20, 200, 400, and 900 U/L) and reasonable recoveries (70%~107%) were obtained. PMID:27240366

  14. Comparison of Prothrombin Time and Aspartate Aminotransferase in Predicting Hepatotoxicity After Acetaminophen Overdose.

    PubMed

    Levine, Michael; O'Connor, Ayrn D; Padilla-Jones, Angela; Gerkin, Richard D

    2016-03-01

    Despite decades of experience with acetaminophen (APAP) overdoses, it remains unclear whether elevated hepatic transaminases or coagulopathy develop first. Furthermore, comparison of the predictive value of these two variables in determining hepatic toxicity following APAP overdoses has been poorly elucidated. The primary objective of this study is to determine the test characteristics of the aspartate aminotransferase (AST) and the prothrombin time (PT) in patients with APAP toxicity. A retrospective chart review of APAP overdoses treated with IV N-acetylcysteine at a tertiary care referral center was performed. Of the 304 subjects included in the study, 246 with an initial AST less than 1000 were analyzed to determine predictors of hepatic injury, defined as an AST exceeding 1000 IU/L. The initial AST >50 was 79.5 % sensitive and 82.6 % specific for predicting hepatic injury. The corresponding negative and positive predictive values were 95.5 and 46.3 %, respectively. In contrast, an initial abnormal PT had a sensitivity of 82.1 % and a specificity of 63.6 %. The negative and positive predictive values for initial PT were 94.9 and 30.2 %, respectively. Although the two tests performed similarly for predicting a composite endpoint of death or liver transplant, neither was a useful predictor. Initial AST performed better than the initial PT for predicting hepatic injury in this series of patients with APAP overdose.

  15. Functional analysis of all aminotransferase proteins inferred from the genome sequence of Corynebacterium glutamicum.

    PubMed

    Marienhagen, Jan; Kennerknecht, Nicole; Sahm, Hermann; Eggeling, Lothar

    2005-11-01

    Twenty putative aminotransferase (AT) proteins of Corynebacterium glutamicum, or rather pyridoxal-5'-phosphate (PLP)-dependent enzymes, were isolated and assayed among others with L-glutamate, L-aspartate, and L-alanine as amino donors and a number of 2-oxo-acids as amino acceptors. One outstanding AT identified is AlaT, which has a broad amino donor specificity utilizing (in the order of preference) L-glutamate > 2-aminobutyrate > L-aspartate with pyruvate as acceptor. Another AT is AvtA, which utilizes L-alanine to aminate 2-oxo-isovalerate, the L-valine precursor, and 2-oxo-butyrate. A second AT active with the L-valine precursor and that of the other two branched-chain amino acids, too, is IlvE, and both enzyme activities overlap partially in vivo, as demonstrated by the analysis of deletion mutants. Also identified was AroT, the aromatic AT, and this and IlvE were shown to have comparable activities with phenylpyruvate, thus demonstrating the relevance of both ATs for L-phenylalanine synthesis. We also assessed the activity of two PLP-containing cysteine desulfurases, supplying a persulfide intermediate. One of them is SufS, which assists in the sulfur transfer pathway for the Fe-S cluster assembly. Together with the identification of further ATs and the additional analysis of deletion mutants, this results in an overview of the ATs within an organism that may not have been achieved thus far.

  16. Histological and Clinical Characteristics of Patients with Chronic Hepatitis C and Persistently Normal Alanine Aminotransferase Levels

    PubMed Central

    Guzman, Grace

    2014-01-01

    Patients with chronic hepatitis C virus (HCV) infection and persistently normal alanine aminotransferase (PNALT) are generally described to have mild liver disease. The aim of this study was to compare clinical and histological features in HCV-infected patients with PNALT and elevated ALT. Patients presenting to the University of Illinois Medical Center, Chicago, who had biopsy proven HCV, an ALT measurement at the time of liver biopsy, at least one additional ALT measurement over the next 12 months, and liver biopsy slides available for review were identified. PNALT was defined as ALT ≤ 30 on at least 2 different occasions over 12 months. Of 1200 patients with HCV, 243 met the study criteria. 13% (32/243) of patients had PNALT while 87% (211/243) had elevated ALT. Significantly more patients with PNALT had advanced fibrosis (F3 and F4) compared to those with elevated ALT (P = 0.007). There was no significant difference in the histology activity index score as well as mean inflammatory score between the two groups. In conclusion, in a well-characterized cohort of patients at a tertiary medical center, PNALT did not distinguish patients with mild liver disease. PMID:24891947

  17. Distribution of D-3-aminoisobutyrate-pyruvate aminotransferase in the rat brain

    PubMed Central

    2014-01-01

    Background D-3-aminoisobutyrate, an intermediary product of thymine, is converted to 2-methyl-3-oxopropanoate using pyruvate as an amino acceptor by D-3-aminoisobutyrate-pyruvate aminotransferase (D-AIB AT; EC 2.6.1.40). A large amount of D-AIB AT is distributed in the kidney and liver; however, small amounts are found in the brain. Recently, D-AIB AT was reported to metabolize asymmetric dimethylarginine (ADMA) in vivo and was suggested to be an important enzyme for nitric oxide metabolism because ADMA is a competitive inhibitor for nitric oxide synthase. In this study, we examined the distribution of D-AIB AT in the rat brain further to understand its role. We measured D-AIB AT mRNA and protein expression using quantitative RT-PCR and Western blotting, and monitored its distribution using immunohistochemical staining. Results D-AIB AT was distributed throughout the brain, with high expression in the cortex and hippocampus. Immunohistochemical staining revealed that D-AIB AT was highly expressed in the retrosplenial cortex and in hippocampal neurons. Conclusion Our results suggest that D-AIB AT is distributed in the examined- just the regions and may play an important role there. PMID:24766736

  18. Aminotransferase levels in relation to short-term use of acetaminophen four grams daily in postoperative cardiothoracic patients in the intensive care unit.

    PubMed

    Ahlers, S J G M; Van Gulik, L; Van Dongen, E P A; Bruins, P; Tibboel, D; Knibbe, C A J

    2011-11-01

    A volunteer study suggested that taking paracetamol 4 g daily could result in elevated alanine aminotransferase plasma levels in a substantial proportion of healthy volunteers. The safety of this dose of paracetamol for acute postoperative pain remains controversial. This study aimed to examine the incidence of alanine aminotransferase elevations after short-term use of paracetamol 4 g daily, as part of the standard pain management protocol, for 93 consecutive patients after cardiothoracic surgery. Alanine aminotransferase levels and other liver function tests were measured preoperatively as baseline and once daily after surgery during the intensive care unit stay. Preoperative alanine aminotransferase levels of more than one time the upper limit of normal (ULN >40 U/l) was observed in 11% (n=10) of the patients but none of these baseline alanine aminotransferase levels exceeded three times the ULN (>3 x ULN). The average daily dose of paracetamol administered was 50 mg/kg (SD=16) after surgery. Postoperative alanine aminotransferase levels of >1 x ULN was observed in 17% (n=16), and 4% (n=4) exceeded >3 x ULN The other liver function tests of the latter four patients, including aspartate aminotransferase (range 173 to 5590 U/l), gamma-glutamyltransferase (range 56 to 103 U/l), lactate dehydrogenase (range 376 to 3518 U/l) and the International Normalised Ratio (range 2.0 to 6.6), were all abnormal. These four patients all had right ventricular failure or cardiogenic shock during the postoperative period which could explain the significant rises in alanine aminotransferase after surgery. In conclusion, the incidence of significant alanine aminotransferase elevations after using daily paracetamol as an analgesic agent for cardiac surgery, at a dose of 4 g per day, was low and mostly due to complications after surgery. Our results, albeit still very limited, provided some reassurance about the safety of paracetamol 4 g daily, as a supplementary analgesic agent for

  19. Structural characterization of AtmS13, a putative sugar aminotransferase involved in indolocarbazole AT2433 aminopentose biosynthesis.

    PubMed

    Singh, Shanteri; Kim, Youngchang; Wang, Fengbin; Bigelow, Lance; Endres, Michael; Kharel, Madan K; Babnigg, Gyorgy; Bingman, Craig A; Joachimiak, Andrzej; Thorson, Jon S; Phillips, George N

    2015-08-01

    AT2433 from Actinomadura melliaura is an indolocarbazole antitumor antibiotic structurally distinguished by its unique aminodideoxypentose-containing disaccharide moiety. The corresponding sugar nucleotide-based biosynthetic pathway for this unusual sugar derives from comparative genomics where AtmS13 has been suggested as the contributing sugar aminotransferase (SAT). Determination of the AtmS13 X-ray structure at 1.50-Å resolution reveals it as a member of the aspartate aminotransferase fold type I (AAT-I). Structural comparisons of AtmS13 with homologous SATs that act upon similar substrates implicate potential active site residues that contribute to distinctions in sugar C5 (hexose vs. pentose) and/or sugar C2 (deoxy vs. hydroxyl) substrate specificity.

  20. Deciphering the Role of Aspartate and Prephenate Aminotransferase Activities in Plastid Nitrogen Metabolism1[C][W][OPEN

    PubMed Central

    de la Torre, Fernando; El-Azaz, Jorge; Ávila, Concepción; Cánovas, Francisco M.

    2014-01-01

    Chloroplasts and plastids of nonphotosynthetic plant cells contain two aspartate (Asp) aminotransferases: a eukaryotic type (Asp5) and a prokaryotic-type bifunctional enzyme displaying Asp and prephenate aminotransferase activities (PAT). We have identified the entire Asp aminotransferase gene family in Nicotiana benthamiana and isolated and cloned the genes encoding the isoenzymes with plastidic localization: NbAsp5 and NbPAT. Using a virus-induced gene silencing approach, we obtained N. benthamiana plants silenced for NbAsp5 and/or NbPAT. Phenotypic and metabolic analyses were conducted in silenced plants to investigate the specific roles of these enzymes in the biosynthesis of essential amino acids within the plastid. The NbAsp5 silenced plants had no changes in phenotype, exhibiting similar levels of free Asp and glutamate as control plants, but contained diminished levels of asparagine and much higher levels of lysine. In contrast, the suppression of NbPAT led to a severe reduction in growth and strong chlorosis symptoms. NbPAT silenced plants exhibited extremely reduced levels of asparagine and were greatly affected in their phenylalanine metabolism and lignin deposition. Furthermore, NbPAT suppression triggered a transcriptional reprogramming in plastid nitrogen metabolism. Taken together, our results indicate that NbPAT has an overlapping role with NbAsp5 in the biosynthesis of Asp and a key role in the production of phenylalanine for the biosynthesis of phenylpropanoids. The analysis of NbAsp5/NbPAT cosilenced plants highlights the central role of both plastidic aminotransferases in nitrogen metabolism; however, only NbPAT is essential for plant growth and development. PMID:24296073

  1. Comparison of blood aminotransferase methods for assessment of myopathy and hepatopathy in Florida manatees (Trichechus manatus latirostris).

    PubMed

    Harr, Kendal E; Allison, Kathryn; Bonde, Robert K; Murphy, David; Harvey, John W

    2008-06-01

    Muscle injury is common in Florida manatees (Trichechus manatus latirostris). Plasma aspartate aminotransferase (AST) is frequently used to assess muscular damage in capture myopathy and traumatic injury. Therefore, accurate measurement of AST and alanine aminotransferase (ALT) is important in managed, free-ranging animals, as well as in those rehabilitating from injury. Activities of these enzymes, however, are usually not increased in manatees with either acute or chronic muscle damage, despite marked increases in plasma creatine kinase activity. It is hypothesized that this absence of response is due to apoenzymes in the blood not detected by commonly used veterinary assays. Addition of coenzyme pyridoxal-5-phosphate (P5P or vitamin B6) should, therefore, result in higher measured enzyme activities. The objective of this study was to determine the most accurate, precise, and diagnostically useful method for aminotransferase measurement in manatees that can be used in veterinary practices and diagnostic laboratories. Additionally, appropriate collection and storage techniques were assessed. The use of an optimized commercial wet chemical assay with 100 micromol P5P resulted in a positive bias of measured enzyme activities in a healthy population of animals. However, AST and ALT were still much lower than that typically observed in domestic animals and should not be used alone in the assessment of capture myopathy and muscular trauma. Additionally, the dry chemistry analyzer, typically used in clinics, reported significantly higher and less precise AST and ALT activities with poor correlation to those measured with wet chemical methods found in diagnostic laboratories. Therefore, these results cannot be clinically compared. Overall, the optimized wet chemical method was the most precise and diagnostically useful measurement of aminotransferase in samples. Additionally, there was a statistically significant difference between paired serum and plasma measurement

  2. [Association between occupational stress and aminotransferase activity in patients with metabolic syndrome].

    PubMed

    Zhao, H; Song, L; Qiang, Y; Liu, H R; Qiu, F Y; Li, X Z; Song, H

    2016-12-20

    Objective: To investigate the association between occupational stress and activity of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in patients with metabolic syndrome. Methods: A case-control study was performed. According to inclusion and exclusion criteria, among the staff members of enterprises and public institutions aged 20~60 years who underwent physical examination in The Affiliated Hospital of Ningxia Medical University and The People's Hospital of Wuzhong from October 2011 to October 2012, 622 patients with metabolic syndrome who did not have a blood relationship with each other were enrolled as case group, and 600 healthy staff members who also did not have a blood relationshipwith each otherwere enrolled as control group. Questionnaire investigation, chronic occupational stress investigation, physical examination, and laboratory tests were performed for all subjects. Results: Compared with the control group, the case group had significantly higher serum levels and abnormal rates of AST and ALT (t=-4.338 and-5.485, χ(2)=11.168 and 34.302, all P<0.05) . There were no significantdifferences in the serum level and abnormal rate of AST between the subgroups with different occupational stresses in both groups (F=2.192 and 2.567, χ(2)=2.694 and 5.402, all P>0.05) , but there were significant differencesbetween the subgroups in all subjects (F=5.005, χ(2)=6.398, all P<0.05) . There were no significant differences in the serum level and abnormal rate of ALT between thesubgroups with different occupational stresses in the case group, the control group, and all subjects (F=0.845, 0.450, and 1.416, χ(2)=2.564, 1.344, and 3.147, all P>0.05) . The partial correlation analysis showed that the total score of occupational stress was positively correlated withthe serum level of AST (r=0.071, P<0.05) and was not correlated with the serum level of ALT (r=-0.044, P>0.05) , and that the serum level of AST was positively correlated with that of ALT

  3. THE ULTRASTRUCTURAL LOCALIZATION OF THE ISOZYMES OF ASPARTATE AMINOTRANSFERASE IN MURINE TISSUES

    PubMed Central

    Papadimitriou, J. M.; Van Duijn, P.

    1970-01-01

    Two isozymes of aspartate aminotransferase have been demonstrated biochemically. One isozyme is found in the mitochondrial fraction of the cytoplasm, the other ("soluble") in the supernatant. Both isozymes can be demonstrated by the cytochemical technique of Lee and Torack, as reported in the preceding report. Aldehyde fixation rapidly inactivates both isozymes, especially the soluble one. Inactivation can be delayed by addition of ketoglutarate to the fixative. The ketoglutarate probably competes with the fixative for the active site of the enzyme, thus protecting that region of the molecule. This enables adequate tissue preservation with enough remaining enzymatic activity to be demonstrated by the precipitation of oxaloacetate as the lead salt from a medium containing α-ketoglutaric acid aspartic acid, and lead nitrate. Electron-opaque material was found not only in mitochondria but, as the result of substrate protection, on the plasma membranes of many cells including erythrocytes and bacteria, the limiting membrane of peroxisomes, and the transverse tubular system of striated muscle. Occasional centrioles, neurotubules, tubules in the tails of spermatozoa, the A-I band junction in myofibrils of striated muscle, and the ground substance between cisternae of endoplasmic reticulum in intestinal goblet cells also showed precipitate. In all cases, replacement of L-aspartic acid by D-aspartic acid in the medium resulted in unstained sections. The sensitivity of extramitochondrial sites to fixation, the need of ketoglutarate as an agent for protecting the enzymatic activity during the fixation process, and the known presence of only soluble isozyme in erythrocytes indicate that enzymatic activity at these sites can be attributed to the soluble isozyme. Localization of the soluble isozyme on the plasma membrane may be related to possible involvement in depolarization phenomena, amino acid transport, or synthesis of plasma membrane-bound mucopolysaccharides. PMID

  4. L,L-Diaminopimelate Aminotransferase from Chlamydomonas reinhardtii: A Target for Algaecide Development

    PubMed Central

    Dobson, Renwick C. J.; Girón, Irma; Hudson, André O.

    2011-01-01

    In some bacterial species and photosynthetic cohorts, including algae, the enzyme l,l-diaminopimelate aminotransferase (DapL) (E.C. 2.6.1.83) is involved in the anabolism of the essential amino acid L-lysine. DapL catalyzes the conversion of tetrahydrodipicolinate (THDPA) to l,l-diaminopimelate (l,l-DAP), in one step bypassing the DapD, DapC and DapE enzymatic reactions present in the acyl DAP pathways. Here we present an in vivo and in vitro characterization of the DapL ortholog from the alga Chlamydomonas reinhardtii (Cr-DapL). The in vivo analysis illustrated that the enzyme is able to functionally complement the E. coli dap auxotrophs and was essential for plant development in Arabidopsis. In vitro, the enzyme was able to inter-convert THDPA and l,l-DAP, showing strong substrate specificity. Cr-DapL was dimeric in both solution and when crystallized. The structure of Cr-DapL was solved in its apo form, showing an overall architecture of a α/β protein with each monomer in the dimer adopting a pyridoxal phosphate-dependent transferase-like fold in a V-shaped conformation. The active site comprises residues from both monomers in the dimer and shows some rearrangement when compared to the apo-DapL structure from Arabidopsis. Since animals do not possess the enzymatic machinery necessary for the de novo synthesis of the amino acid l-lysine, enzymes involved in this pathway are attractive targets for the development of antibiotics, herbicides and algaecides. PMID:21633707

  5. L,L-diaminopimelate aminotransferase from Chlamydomonas reinhardtii: a target for algaecide development.

    PubMed

    Dobson, Renwick C J; Girón, Irma; Hudson, André O

    2011-01-01

    In some bacterial species and photosynthetic cohorts, including algae, the enzyme L,L-diaminopimelate aminotransferase (DapL) (E.C. 2.6.1.83) is involved in the anabolism of the essential amino acid L-lysine. DapL catalyzes the conversion of tetrahydrodipicolinate (THDPA) to L,L-diaminopimelate (L,L-DAP), in one step bypassing the DapD, DapC and DapE enzymatic reactions present in the acyl DAP pathways. Here we present an in vivo and in vitro characterization of the DapL ortholog from the alga Chlamydomonas reinhardtii (Cr-DapL). The in vivo analysis illustrated that the enzyme is able to functionally complement the E. coli dap auxotrophs and was essential for plant development in Arabidopsis. In vitro, the enzyme was able to inter-convert THDPA and L,L-DAP, showing strong substrate specificity. Cr-DapL was dimeric in both solution and when crystallized. The structure of Cr-DapL was solved in its apo form, showing an overall architecture of a α/β protein with each monomer in the dimer adopting a pyridoxal phosphate-dependent transferase-like fold in a V-shaped conformation. The active site comprises residues from both monomers in the dimer and shows some rearrangement when compared to the apo-DapL structure from Arabidopsis. Since animals do not possess the enzymatic machinery necessary for the de novo synthesis of the amino acid L-lysine, enzymes involved in this pathway are attractive targets for the development of antibiotics, herbicides and algaecides.

  6. Inhibition of the enzyme, GABA-aminotransferase in human platelets by vigabatrin, a potential antiepileptic drug.

    PubMed Central

    Rimmer, E; Kongola, G; Richens, A

    1988-01-01

    1. The effect of the new antiepileptic drug, vigabatrin (gamma-vinyl GABA), on the platelet enzyme, GABA-aminotransferase (GABA-T) was investigated in volunteers and patients. Platelets GABA-T activity was assayed using a radioenzymic method. 2. Three single oral doses of vigabatrin (1 g, 2 g and 4 g) were given to six healthy male volunteers in an open randomised cross over study and compared with a baseline period preceding the three treatments. 3. Significant inhibition of the platelet GABA-T was produced by treatment with all three doses and a dose-response relationship was demonstrated. The minimum enzyme activities after 1 g, 2 g and 4 g doses were 43%, 30% and 21% respectively compared with the control values. 4. A significant depression of enzyme activity occurred at 30 min after drug administration and the values remained below control values for 72 h post-dose, outlasting the presence of the drug itself in the plasma. 5. Eight patients with chronic refractory epilepsy were treated with vigabatrin for 6 weeks. After taking the 2 g daily dose for 1 week there was a marked reduction in platelet enzyme activity in all subjects but the enzyme inhibition produced by the 3 g dose was not significantly different from that produced by the 2 g dose, even after 4 weeks treatment with the larger dose. The mean enzyme activity was approximately 30% throughout the active treatment period. One week after stopping vigabatrin, the enzyme levels were not significantly different from the baseline values.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:3358887

  7. Association between Serum Uric Acid and Elevated Alanine Aminotransferase in the General Population

    PubMed Central

    Chen, Shuang; Guo, Xiaofan; Yu, Shasha; Sun, Guozhe; Yang, Hongmei; Li, Zhao; Sun, Yingxian

    2016-01-01

    Background: Both the serum uric acid (SUA) level and elevated alanine aminotransferase (ALT) are related to metabolic syndrome. However, the association between SUA and elevated ALT has not been elucidated in the general population. The objective of this study was to investigate the association between SUA and elevated ALT in the general population of China; Methods: A total of 11,572 adults (≥35 years of age) participated in this survey. Elevated ALT was defined as >40 U/L. SUA ≥ 7.0 mg/dL in males or ≥6.0 mg/dL in females was defined as hyperuricemia. SUA within the reference range was divided into quartiles, and its associations with elevated ALT were evaluated by logistic regressions; Results: A total of 7.4% participants had elevated ALT. The prevalence of hyperuricemia was 14.9% in males and 7.3% in females. There was a significantly positive dose-response association between SUA levels and the prevalence of elevated ALT. After adjusting for potential confounders, a positive relationship for elevated ALT was observed in subjects with hyperuricemia (odds ratio [OR]: 2.032, 95% confidence interval [CI]: 1.443–2.861 for men; OR: 2.045, 95% CI: 1.221–3.425 for women, both p < 0.05). Within the reference range, the association between SUA and elevated ALT persisted in the fourth quartile (OR: 1.467, 95% CI: 1.063–2.025 for men; OR: 1.721, 95% CI: 1.146–2.585 for women, both p < 0.05); Conclusions: Our results indicated that an increased SUA level, even within the reference range, was independently associated with elevated ALT in Chinese adults. PMID:27563918

  8. Characterization of kynurenine aminotransferase III, a novel member of a phylogenetically conserved KAT family.

    PubMed

    Yu, Ping; Li, Zhengsheng; Zhang, Ling; Tagle, Danilo A; Cai, Tao

    2006-01-03

    Kynurenine aminotransferase (KAT) is an enzyme responsible for synthesis of kynurenic acid (KYNA), a well established neuroprotective and anticonvulsant agent, involved in synaptic transmission and implicated in the pathophysiology of schizophrenia, Huntington's disease and other neurological disorders. We have shown previously that kat2-/- mice had lower hippocampal KYNA levels and were more hyperactive than wild-type mice. However, these abnormalities occur early and are transitory coinciding with restoration of KYNA levels, suggesting that compensatory changes or ontogenetic expression of another unknown homolog may account for the normalization of KYNA levels in the adult kat2-/- mice brain. Here, we report the isolation of a novel KAT molecule, kat3, from mouse and human brain cDNA libraries. The encoded 454 amino acids of human KAT III share 64.8% similarity to that of KAT I and 30.1% to KAT II. Northern blot analysis demonstrated that kat3 mRNA is widely expressed but with higher expression levels in liver, kidney, heart, and neuroendocrine tissues. RT-PCR and Northern analysis showed that kat3 expression starts as early as postnatal day (PND) 7 and peaks in adult. The mRNA level of kat3 and kat1 when measured together is significantly higher at PND 60 in kat2-/- mice than those of wild-type mice indicating possible co-regulation of expression levels. RNA-interference (RNAi) directed towards transcripts for either R03A10.4 or F28H6.3 in Caenorhabditis elegans which are kat1 and kat3 orthologs, respectively, did not result in any gross abnormalities. Our results show that upregulation of kat3 and kat1 may be responsible for the phenotypic rescue on kat2-/- mice.

  9. Biochemical and phenotypic abnormalities in kynurenine aminotransferase II-deficient mice.

    PubMed

    Yu, Ping; Di Prospero, Nicholas A; Sapko, Michael T; Cai, Tao; Chen, Amy; Melendez-Ferro, Miguel; Du, Fu; Whetsell, William O; Guidetti, Paolo; Schwarcz, Robert; Tagle, Danilo A

    2004-08-01

    Kynurenic acid (KYNA) can act as an endogenous modulator of excitatory neurotransmission and has been implicated in the pathogenesis of several neurological and psychiatric diseases. To evaluate its role in the brain, we disrupted the murine gene for kynurenine aminotransferase II (KAT II), the principal enzyme responsible for the synthesis of KYNA in the rat brain. mKat-2(-/-) mice showed no detectable KAT II mRNA or protein. Total brain KAT activity and KYNA levels were reduced during the first month but returned to normal levels thereafter. In contrast, liver KAT activity and KYNA levels in mKat-2(-/-) mice were decreased by >90% throughout life, though no hepatic abnormalities were observed histologically. KYNA-associated metabolites kynurenine, 3-hydroxykynurenine, and quinolinic acid were unchanged in the brain and liver of knockout mice. mKat-2(-/-) mice began to manifest hyperactivity and abnormal motor coordination at 2 weeks of age but were indistinguishable from wild type after 1 month of age. Golgi staining of cortical and striatal neurons revealed enlarged dendritic spines and a significant increase in spine density in 3-week-old mKat-2(-/-) mice but not in 2-month-old animals. Our results show that gene targeting of mKat-2 in mice leads to early and transitory decreases in brain KAT activity and KYNA levels with commensurate behavioral and neuropathological changes and suggest that compensatory changes or ontogenic expression of another isoform may account for the normalization of KYNA levels in the adult mKat-2(-/-) brain.

  10. Auxin and Tryptophan Homeostasis Are Facilitated by the ISS1/VAS1 Aromatic Aminotransferase in Arabidopsis

    PubMed Central

    Pieck, Michael; Yuan, Youxi; Godfrey, Jason; Fisher, Christopher; Zolj, Sanda; Vaughan, Dylan; Thomas, Nicholas; Wu, Connie; Ramos, Julian; Lee, Norman; Normanly, Jennifer; Celenza, John L.

    2015-01-01

    Indole-3-acetic acid (IAA) plays a critical role in regulating numerous aspects of plant growth and development. While there is much genetic support for tryptophan-dependent (Trp-D) IAA synthesis pathways, there is little genetic evidence for tryptophan-independent (Trp-I) IAA synthesis pathways. Using Arabidopsis, we identified two mutant alleles of ISS1 (Indole Severe Sensitive) that display indole-dependent IAA overproduction phenotypes including leaf epinasty and adventitious rooting. Stable isotope labeling showed that iss1, but not WT, uses primarily Trp-I IAA synthesis when grown on indole-supplemented medium. In contrast, both iss1 and WT use primarily Trp-D IAA synthesis when grown on unsupplemented medium. iss1 seedlings produce 8-fold higher levels of IAA when grown on indole and surprisingly have a 174-fold increase in Trp. These findings indicate that the iss1 mutant’s increase in Trp-I IAA synthesis is due to a loss of Trp catabolism. ISS1 was identified as At1g80360, a predicted aromatic aminotransferase, and in vitro and in vivo analysis confirmed this activity. At1g80360 was previously shown to primarily carry out the conversion of indole-3-pyruvic acid to Trp as an IAA homeostatic mechanism in young seedlings. Our results suggest that in addition to this activity, in more mature plants ISS1 has a role in Trp catabolism and possibly in the metabolism of other aromatic amino acids. We postulate that this loss of Trp catabolism impacts the use of Trp-D and/or Trp-I IAA synthesis pathways. PMID:26163189

  11. Association between Elevated Alanine Aminotransferase and Urosepsis in Children with Acute Pyelonephritis

    PubMed Central

    Kim, Dongwan; Lee, Sung Hyun; Ryoo, Eell; Cho, Hye Kyung; Kim, Yun Mi

    2016-01-01

    Purpose The aim of this study is to investigate the association between elevated alanine aminotransferase (ALT) and urosepsis in children with acute pyelonephritis (APN). Methods We retrospectively identified all children who were managed in our hospital with APN during a decade period. In our study a diagnosis of APN was defined as having a positive urine culture and a positive (99m)Tc-dimercaptosuccinic acid scintigraphy. We compared those with elevated ALT and those with normal ALT according to the following variables: age, gender, duration of fever prior to admission, presence of hypotension, C-reactive protein (CRP), creatinine, presence of anemia, white blood cells count, platelet count, blood culture result, and grades of vesicoureteral reflux. In addition, the correlation between elevated ALT and positive blood culture was analyzed in detail. Results A total of 996 children were diagnosed with APN, of which 883 were included in the study. ALT was elevated in 81 children (9.2%). In the analysis of demographic characteristics, the number of children with elevated ALT was higher in children between 0 to 3 months, boys, and in those with positive blood culture (p=0.002, 0.036, and 0.010, respectively). In multivariate analysis of variables associated with positive blood culture, age younger than 3 months, elevated ALT, elevated CRP, and elevated creatinine showed statistical significance (p=0.004, 0.030, 0.043, and 0.044, respectively). Conclusion Our study demonstrates the association between elevated ALT and increased prevalence of urosepsis in addition to elevated CRP, elevated creatinine, and age younger than 3 months in children with APN. PMID:27066449

  12. Functional Analysis of All Aminotransferase Proteins Inferred from the Genome Sequence of Corynebacterium glutamicum

    PubMed Central

    Marienhagen, Jan; Kennerknecht, Nicole; Sahm, Hermann; Eggeling, Lothar

    2005-01-01

    Twenty putative aminotransferase (AT) proteins of Corynebacterium glutamicum, or rather pyridoxal-5′-phosphate (PLP)-dependent enzymes, were isolated and assayed among others with l-glutamate, l-aspartate, and l-alanine as amino donors and a number of 2-oxo-acids as amino acceptors. One outstanding AT identified is AlaT, which has a broad amino donor specificity utilizing (in the order of preference) l-glutamate > 2-aminobutyrate > l-aspartate with pyruvate as acceptor. Another AT is AvtA, which utilizes l-alanine to aminate 2-oxo-isovalerate, the l-valine precursor, and 2-oxo-butyrate. A second AT active with the l-valine precursor and that of the other two branched-chain amino acids, too, is IlvE, and both enzyme activities overlap partially in vivo, as demonstrated by the analysis of deletion mutants. Also identified was AroT, the aromatic AT, and this and IlvE were shown to have comparable activities with phenylpyruvate, thus demonstrating the relevance of both ATs for l-phenylalanine synthesis. We also assessed the activity of two PLP-containing cysteine desulfurases, supplying a persulfide intermediate. One of them is SufS, which assists in the sulfur transfer pathway for the Fe-S cluster assembly. Together with the identification of further ATs and the additional analysis of deletion mutants, this results in an overview of the ATs within an organism that may not have been achieved thus far. PMID:16267288

  13. Relaxed Evolution in the Tyrosine Aminotransferase Gene Tat in Old World Fruit Bats (Chiroptera: Pteropodidae)

    PubMed Central

    Shen, Bin; Fang, Tao; Yang, Tianxiao; Jones, Gareth; Irwin, David M.; Zhang, Shuyi

    2014-01-01

    Frugivorous and nectarivorous bats fuel their metabolism mostly by using carbohydrates and allocate the restricted amounts of ingested proteins mainly for anabolic protein syntheses rather than for catabolic energy production. Thus, it is possible that genes involved in protein (amino acid) catabolism may have undergone relaxed evolution in these fruit- and nectar-eating bats. The tyrosine aminotransferase (TAT, encoded by the Tat gene) is the rate-limiting enzyme in the tyrosine catabolic pathway. To test whether the Tat gene has undergone relaxed evolution in the fruit- and nectar-eating bats, we obtained the Tat coding region from 20 bat species including four Old World fruit bats (Pteropodidae) and two New World fruit bats (Phyllostomidae). Phylogenetic reconstructions revealed a gene tree in which all echolocating bats (including the New World fruit bats) formed a monophyletic group. The phylogenetic conflict appears to stem from accelerated TAT protein sequence evolution in the Old World fruit bats. Our molecular evolutionary analyses confirmed a change in the selection pressure acting on Tat, which was likely caused by a relaxation of the evolutionary constraints on the Tat gene in the Old World fruit bats. Hepatic TAT activity assays showed that TAT activities in species of the Old World fruit bats are significantly lower than those of insectivorous bats and omnivorous mice, which was not caused by a change in TAT protein levels in the liver. Our study provides unambiguous evidence that the Tat gene has undergone relaxed evolution in the Old World fruit bats in response to changes in their metabolism due to the evolution of their special diet. PMID:24824435

  14. Tyrosine Aminotransferase Contributes to Benzylisoquinoline Alkaloid Biosynthesis in Opium Poppy1[W

    PubMed Central

    Lee, Eun-Jeong; Facchini, Peter J.

    2011-01-01

    Tyrosine aminotransferase (TyrAT) catalyzes the transamination of l-Tyr and α-ketoglutarate, yielding 4-hydroxyphenylpyruvic acid and l-glutamate. The decarboxylation product of 4-hydroxyphenylpyruvic acid, 4-hydroxyphenylacetaldehyde, is a precursor to a large and diverse group of natural products known collectively as benzylisoquinoline alkaloids (BIAs). We have isolated and characterized a TyrAT cDNA from opium poppy (Papaver somniferum), which remains the only commercial source for several pharmaceutical BIAs, including codeine, morphine, and noscapine. TyrAT belongs to group I pyridoxal 5′-phosphate (PLP)-dependent enzymes wherein Schiff base formation occurs between PLP and a specific Lys residue. The amino acid sequence of TyrAT showed considerable homology to other putative plant TyrATs, although few of these have been functionally characterized. Purified, recombinant TyrAT displayed a molecular mass of approximately 46 kD and a substrate preference for l-Tyr and α-ketoglutarate, with apparent Km values of 1.82 and 0.35 mm, respectively. No specific requirement for PLP was detected in vitro. Liquid chromatography-tandem mass spectrometry confirmed the conversion of l-Tyr to 4-hydroxyphenylpyruvate. TyrAT gene transcripts were most abundant in roots and stems of mature opium poppy plants. Virus-induced gene silencing was used to evaluate the contribution of TyrAT to BIA metabolism in opium poppy. TyrAT transcript levels were reduced by at least 80% in silenced plants compared with controls and showed a moderate reduction in total alkaloid content. The modest correlation between transcript levels and BIA accumulation in opium poppy supports a role for TyrAT in the generation of alkaloid precursors, but it also suggests the occurrence of other sources for 4-hydroxyphenylacetaldehyde. PMID:21949209

  15. Branched-Chain Aminotransferases Control TORC1 Signaling in Saccharomyces cerevisiae.

    PubMed

    Kingsbury, Joanne M; Sen, Neelam D; Cardenas, Maria E

    2015-12-01

    The conserved target of rapamycin complex 1 (TORC1) integrates nutrient signals to orchestrate cell growth and proliferation. Leucine availability is conveyed to control TORC1 activity via the leu-tRNA synthetase/EGOC-GTPase module in yeast and mammals, but the mechanisms sensing leucine remain only partially understood. We show here that both leucine and its α-ketoacid metabolite, α-ketoisocaproate, effectively activate the yeast TORC1 kinase via both EGOC GTPase-dependent and -independent mechanisms. Leucine and α-ketoisocaproate are interconverted by ubiquitous branched-chain aminotransferases (BCAT), which in yeast are represented by the mitochondrial and cytosolic enzymes Bat1 and Bat2, respectively. BCAT yeast mutants exhibit severely compromised TORC1 activity, which is partially restored by expression of Bat1 active site mutants, implicating both catalytic and structural roles of BCATs in TORC1 control. We find that Bat1 interacts with branched-chain amino acid metabolic enzymes and, in a leucine-dependent fashion, with the tricarboxylic acid (TCA)-cycle enzyme aconitase. BCAT mutation perturbed TCA-cycle intermediate levels, consistent with a TCA-cycle block, and resulted in low ATP levels, activation of AMPK, and TORC1 inhibition. We propose the biosynthetic capacity of BCAT and its role in forming multicomplex metabolons connecting branched-chain amino acids and TCA-cycle metabolism governs TCA-cycle flux to activate TORC1 signaling. Because mammalian mitochondrial BCAT is known to form a supramolecular branched-chain α-keto acid dehydrogenase enzyme complex that links leucine metabolism to the TCA-cycle, these findings establish a precedent for understanding TORC1 signaling in mammals.

  16. Altered Expression of Human Mitochondrial Branched Chain Aminotransferase in Dementia with Lewy Bodies and Vascular Dementia.

    PubMed

    Ashby, Emma L; Kierzkowska, Marta; Hull, Jonathon; Kehoe, Patrick G; Hutson, Susan M; Conway, Myra E

    2017-01-01

    Cytosolic and mitochondrial human branched chain aminotransferase (hBCATc and hBCATm, respectively) play an integral role in brain glutamate metabolism. Regional increased levels of hBCATc in the CA1 and CA4 region of Alzheimer's disease (AD) brain together with increased levels of hBCATm in frontal and temporal cortex of AD brains, suggest a role for these proteins in glutamate excitotoxicity. Glutamate toxicity is a key pathogenic feature of several neurological disorders including epilepsy associated dementia, AD, vascular dementia (VaD) and dementia with Lewy bodies (DLB). To further understand if these increases are specific to AD, the expression profiles of hBCATc and hBCATm were examined in other forms of dementia including DLB and VaD. Similar to AD, levels of hBCATm were significantly increased in the frontal and temporal cortex of VaD cases and in frontal cortex of DLB cases compared to controls, however there were no observed differences in hBCATc between groups in these areas. Moreover, multiple forms of hBCATm were observed that were particular to the disease state relative to matched controls. Real-time PCR revealed similar expression of hBCATm mRNA in frontal and temporal cortex for all cohort comparisons, whereas hBCATc mRNA expression was significantly increased in VaD cases compared to controls. Collectively our results suggest that hBCATm protein expression is significantly increased in the brains of DLB and VaD cases, similar to those reported in AD brain. These findings indicate a more global response to altered glutamate metabolism and suggest common metabolic responses that might reflect shared neurodegenerative mechanisms across several forms of dementia.

  17. Branched-Chain Aminotransferases Control TORC1 Signaling in Saccharomyces cerevisiae

    PubMed Central

    Kingsbury, Joanne M.; Sen, Neelam D.; Cardenas, Maria E.

    2015-01-01

    The conserved target of rapamycin complex 1 (TORC1) integrates nutrient signals to orchestrate cell growth and proliferation. Leucine availability is conveyed to control TORC1 activity via the leu-tRNA synthetase/EGOC-GTPase module in yeast and mammals, but the mechanisms sensing leucine remain only partially understood. We show here that both leucine and its α-ketoacid metabolite, α-ketoisocaproate, effectively activate the yeast TORC1 kinase via both EGOC GTPase-dependent and -independent mechanisms. Leucine and α-ketoisocaproate are interconverted by ubiquitous branched-chain aminotransferases (BCAT), which in yeast are represented by the mitochondrial and cytosolic enzymes Bat1 and Bat2, respectively. BCAT yeast mutants exhibit severely compromised TORC1 activity, which is partially restored by expression of Bat1 active site mutants, implicating both catalytic and structural roles of BCATs in TORC1 control. We find that Bat1 interacts with branched-chain amino acid metabolic enzymes and, in a leucine-dependent fashion, with the tricarboxylic acid (TCA)-cycle enzyme aconitase. BCAT mutation perturbed TCA-cycle intermediate levels, consistent with a TCA-cycle block, and resulted in low ATP levels, activation of AMPK, and TORC1 inhibition. We propose the biosynthetic capacity of BCAT and its role in forming multicomplex metabolons connecting branched-chain amino acids and TCA-cycle metabolism governs TCA-cycle flux to activate TORC1 signaling. Because mammalian mitochondrial BCAT is known to form a supramolecular branched-chain α-keto acid dehydrogenase enzyme complex that links leucine metabolism to the TCA-cycle, these findings establish a precedent for understanding TORC1 signaling in mammals. PMID:26659116

  18. Relaxed evolution in the tyrosine aminotransferase gene tat in old world fruit bats (Chiroptera: Pteropodidae).

    PubMed

    Shen, Bin; Fang, Tao; Yang, Tianxiao; Jones, Gareth; Irwin, David M; Zhang, Shuyi

    2014-01-01

    Frugivorous and nectarivorous bats fuel their metabolism mostly by using carbohydrates and allocate the restricted amounts of ingested proteins mainly for anabolic protein syntheses rather than for catabolic energy production. Thus, it is possible that genes involved in protein (amino acid) catabolism may have undergone relaxed evolution in these fruit- and nectar-eating bats. The tyrosine aminotransferase (TAT, encoded by the Tat gene) is the rate-limiting enzyme in the tyrosine catabolic pathway. To test whether the Tat gene has undergone relaxed evolution in the fruit- and nectar-eating bats, we obtained the Tat coding region from 20 bat species including four Old World fruit bats (Pteropodidae) and two New World fruit bats (Phyllostomidae). Phylogenetic reconstructions revealed a gene tree in which all echolocating bats (including the New World fruit bats) formed a monophyletic group. The phylogenetic conflict appears to stem from accelerated TAT protein sequence evolution in the Old World fruit bats. Our molecular evolutionary analyses confirmed a change in the selection pressure acting on Tat, which was likely caused by a relaxation of the evolutionary constraints on the Tat gene in the Old World fruit bats. Hepatic TAT activity assays showed that TAT activities in species of the Old World fruit bats are significantly lower than those of insectivorous bats and omnivorous mice, which was not caused by a change in TAT protein levels in the liver. Our study provides unambiguous evidence that the Tat gene has undergone relaxed evolution in the Old World fruit bats in response to changes in their metabolism due to the evolution of their special diet.

  19. Methanococci use the diaminopimelate aminotransferase (DapL) pathway for lysine biosynthesis.

    PubMed

    Liu, Yuchen; White, Robert H; Whitman, William B

    2010-07-01

    The pathway of lysine biosynthesis in the methanococci has not been identified previously. A variant of the diaminopimelic acid (DAP) pathway uses diaminopimelate aminotransferase (DapL) to catalyze the direct conversion of tetrahydrodipicolinate (THDPA) to ll-DAP. Recently, the enzyme DapL (MTH52) was identified in Methanothermobacter thermautotrophicus and shown to belong to the DapL1 group. Although the Methanococcus maripaludis genome lacks a gene that can be unambiguously assigned a DapL function based on sequence similarity, the open reading frame MMP1527 product shares 30% amino acid sequence identity with MTH52. A Deltammp1527 deletion mutant was constructed and found to be a lysine auxotroph, suggesting that this DapL homolog in methanococci is required for lysine biosynthesis. In cell extracts of the M. maripaludis wild-type strain, the specific activity of DapL using ll-DAP and alpha-ketoglutarate as substrates was 24.3 + or - 2.0 nmol min(-1) mg of protein(-1). The gene encoding the DapL homolog in Methanocaldococcus jannaschii (MJ1391) was cloned and expressed in Escherichia coli, and the protein was purified. The maximum activity of MJ1391 was observed at 70 degrees C and pH 8.0 to 9.0. The apparent K(m)s of MJ1391 for ll-DAP and alpha-ketoglutarate were 82.8 + or - 10 microM and 0.42 + or - 0.02 mM, respectively. MJ1391 was not able to use succinyl-DAP or acetyl-DAP as a substrate. Phylogenetic analyses suggested that two lateral gene transfers occurred in the DapL genes, one from the archaea to the bacteria in the DapL2 group and one from the bacteria to the archaea in the DapL1 group. These results demonstrated that the DapL pathway is present in marine methanogens belonging to the Methanococcales.

  20. Methanococci Use the Diaminopimelate Aminotransferase (DapL) Pathway for Lysine Biosynthesis ▿

    PubMed Central

    Liu, Yuchen; White, Robert H.; Whitman, William B.

    2010-01-01

    The pathway of lysine biosynthesis in the methanococci has not been identified previously. A variant of the diaminopimelic acid (DAP) pathway uses diaminopimelate aminotransferase (DapL) to catalyze the direct conversion of tetrahydrodipicolinate (THDPA) to ll-DAP. Recently, the enzyme DapL (MTH52) was identified in Methanothermobacter thermautotrophicus and shown to belong to the DapL1 group. Although the Methanococcus maripaludis genome lacks a gene that can be unambiguously assigned a DapL function based on sequence similarity, the open reading frame MMP1527 product shares 30% amino acid sequence identity with MTH52. A Δmmp1527 deletion mutant was constructed and found to be a lysine auxotroph, suggesting that this DapL homolog in methanococci is required for lysine biosynthesis. In cell extracts of the M. maripaludis wild-type strain, the specific activity of DapL using ll-DAP and α-ketoglutarate as substrates was 24.3 ± 2.0 nmol min−1 mg of protein−1. The gene encoding the DapL homolog in Methanocaldococcus jannaschii (MJ1391) was cloned and expressed in Escherichia coli, and the protein was purified. The maximum activity of MJ1391 was observed at 70°C and pH 8.0 to 9.0. The apparent Kms of MJ1391 for ll-DAP and α-ketoglutarate were 82.8 ± 10 μM and 0.42 ± 0.02 mM, respectively. MJ1391 was not able to use succinyl-DAP or acetyl-DAP as a substrate. Phylogenetic analyses suggested that two lateral gene transfers occurred in the DapL genes, one from the archaea to the bacteria in the DapL2 group and one from the bacteria to the archaea in the DapL1 group. These results demonstrated that the DapL pathway is present in marine methanogens belonging to the Methanococcales. PMID:20418392

  1. Monitoring of extracellular aspartate aminotransferase and troponin T by microdialysis during and after cardioplegic heart arrest.

    PubMed

    Kennergren, C; Mantovani, V; Lönnroth, P; Nyström, B; Berglin, E; Hamberger, A

    1999-01-01

    This study aims at developing per- and postopertive surveillance of the myocardium and focuses on ischemic damage following cardioplegic heart arrest. Levels of troponin T and total aspartate aminotransferase (ASAT) were analyzed in the myocardial interstitium of 10 patients with ischemic heart disease (IHD) who underwent coronary bypass surgery and in 12 patients with nonischemic heart disease (N-IHD) who underwent valvular surgery. Fluid from the myocardial interstitium of the anterior and the lateral wall of the heart was sampled by microdialysis probes that were implanted during surgery and extracted percutaneously 70-100 h later. There were no adverse reactions, and the equipment did not interfere with the surgical procedures. The peak in troponin T serum levels that occurred 4 h after cardiac arrest was preceded by a peak in troponin T levels in the microdialysates from the interstitium that occurred 1 h earlier. The concentration of troponin T in the microdialysate peak was 300 times higher than in the serum peak. The increase in serum ASAT levels during the first 7 h after cardiac arrest corresponded in time with a decrease in interstitial ASAT levels, which had already reached a maximum during cardiac arrest. The microdialysate/serum concentration ratio was considerably smaller for ASAT than for troponin T. Interstitial peak levels of troponin T correlated positively and significantly with peak levels of ASAT. Of the 22 patients, 15 had no postoperative events according to clinical outcome, ECG and serum tests. Fourteen of these had low to normal levels of interstitial ASAT and troponin T. Conversely, atrial fibrillation and/or premature atrial contractions were recorded in 8/22 patients, 7 of whom had elevated interstitial ASAT and/or troponin T concentrations in one or both of the sampled heart regions. The N-IHD patients had higher levels of troponin T in the interstitium 20-70 h following cardioplegia, while the peak levels did not differ between the

  2. Structural basis of substrate selectivity of Δ(1)-pyrroline-5-carboxylate dehydrogenase (ALDH4A1): semialdehyde chain length.

    PubMed

    Pemberton, Travis A; Tanner, John J

    2013-10-01

    The enzyme Δ(1)-pyrroline-5-carboxylate (P5C) dehydrogenase (aka P5CDH and ALDH4A1) is an aldehyde dehydrogenase that catalyzes the oxidation of γ-glutamate semialdehyde to l-glutamate. The crystal structures of mouse P5CDH complexed with glutarate, succinate, malonate, glyoxylate, and acetate are reported. The structures are used to build a structure-activity relationship that describes the semialdehyde carbon chain length and the position of the aldehyde group in relation to the cysteine nucleophile and oxyanion hole. Efficient 4- and 5-carbon substrates share the common feature of being long enough to span the distance between the anchor loop at the bottom of the active site and the oxyanion hole at the top of the active site. The inactive 2- and 3-carbon semialdehydes bind the anchor loop but are too short to reach the oxyanion hole. Inhibition of P5CDH by glyoxylate, malonate, succinate, glutarate, and l-glutamate is also examined. The Ki values are 0.27 mM for glyoxylate, 58 mM for succinate, 30 mM for glutarate, and 12 mM for l-glutamate. Curiously, malonate is not an inhibitor. The trends in Ki likely reflect a trade-off between the penalty for desolvating the carboxylates of the free inhibitor and the number of compensating hydrogen bonds formed in the enzyme-inhibitor complex.

  3. Reconfiguration of N Metabolism upon Hypoxia Stress and Recovery: Roles of Alanine Aminotransferase (AlaAT) and Glutamate Dehydrogenase (GDH)

    PubMed Central

    Diab, Houssein; Limami, Anis M.

    2016-01-01

    In the context of climatic change, more heavy precipitation and more frequent flooding and waterlogging events threaten the productivity of arable farmland. Furthermore, crops were not selected to cope with flooding- and waterlogging-induced oxygen limitation. In general, low oxygen stress, unlike other abiotic stresses (e.g., cold, high temperature, drought and saline stress), received little interest from the scientific community and less financial support from stakeholders. Accordingly, breeding programs should be developed and agronomical practices should be adapted in order to save plants’ growth and yield—even under conditions of low oxygen availability (e.g., submergence and waterlogging). The prerequisite to the success of such breeding programs and changes in agronomical practices is a good knowledge of how plants adapt to low oxygen stress at the cellular and the whole plant level. In the present paper, we summarized the recent knowledge on metabolic adjustment in general under low oxygen stress and highlighted thereafter the major changes pertaining to the reconfiguration of amino acids syntheses. We propose a model showing (i) how pyruvate derived from active glycolysis upon hypoxia is competitively used by the alanine aminotransferase/glutamate synthase cycle, leading to alanine accumulation and NAD+ regeneration. Carbon is then saved in a nitrogen store instead of being lost through ethanol fermentative pathway. (ii) During the post-hypoxia recovery period, the alanine aminotransferase/glutamate dehydrogenase cycle mobilizes this carbon from alanine store. Pyruvate produced by the reverse reaction of alanine aminotransferase is funneled to the TCA cycle, while deaminating glutamate dehydrogenase regenerates, reducing equivalent (NADH) and 2-oxoglutarate to maintain the cycle function. PMID:27258319

  4. Identification of stsC, the gene encoding the L-glutamine:scyllo-inosose aminotransferase from streptomycin-producing Streptomycetes.

    PubMed

    Ahlert, J; Distler, J; Mansouri, K; Piepersberg, W

    1997-08-01

    Eight new genes, strO-stsABCDEFG, were identified by sequencing DNA in the gene cluster that encodes proteins for streptomycin production of Streptomyces griseus N2-3-11. The StsA (calculated molecular mass 43.5 kDa) and StsC (45.5 kDa) proteins - together with another gene product, StrS (39.8 kDa), encoded in another operon of the same gene cluster - show significant sequence identity and are members of a new class of pyridoxal-phosphate-dependent aminotransferases that have been observed mainly in the biosynthetic pathways for secondary metabolites. The aminotransferase activity was demonstrated for the first time by identification of the overproduced and purified StsC protein as the L-glutamine:scyllo-inosose aminotransferase, which catalyzes the first amino transfer in the biosynthesis of the streptidine subunit of streptomycin. The stsC and stsA genes each hybridized specifically to distinct fragments in the genomic DNA of most actinomycetes tested that produce diaminocyclitolaminoglycosides. In contrast, only stsC, but not stsA, hybridized to the DNA of Streptomyces hygroscopicus ssp. glebosus, which produces the monoaminocyclitol antibiotic bluensomycin; this suggests that both genes are specifically used in the first and second steps of the cyclitol transamination reactions. Sequence comparison studies performed with the deduced polypeptides of the genes adjacent to stsC suggest that the enzymes encoded by some of these genes [strO (putative phosphatase gene), stsB (putative oxidoreductase gene), and stsE (putative phosphotransferase gene)] also could be involved in (di-)aminocyclitol synthesis.

  5. Comparison of blood aminotransferase methods for assessment of myopathy and hepatopathy in Florida manatees (Trichechus manatus latirostris)

    USGS Publications Warehouse

    Harr, K.E.; Allison, K.; Bonde, R.K.; Murphy, D.; Harvey, J.W.

    2008-01-01

    Muscle injury is common in Florida manatees (Trichechus manatus latirostris). Plasma aspartate amino-transferase (AST) is frequently used to assess muscular damage in capture myopathy and traumatic injury. Therefore, accurate measurement of AST and alanine aminotransferase (ALT) is important in managed, free-ranging animals, as well as in those rehabilitating from injury. Activities of these enzymes, however, are usually not increased in manatees with either acute or chronic muscle damage, despite marked increases in plasma creatine kinase activity. It is hypothesized that this absence of response is due to apoenzymes in the blood not detected by commonly used veterinary assays. Addition of coenzyme pyridoxal-5-phosphate (P5P or vitamin B6) should, therefore, result in higher measured enzyme activities. The objective of this study was to determine the most accurate, precise, and diagnostically useful method for aminotransferase measurement in manatees that can be used in veterinary practices and diagnostic laboratories. Additionally, appropriate collection and storage techniques were assessed. The use of an optimized commercial wet chemical assay with 100 ??mol P5P resulted in a positive bias of measured enzyme activities in a healthy population of animals. However, AST and ALT were still much lower than that typically observed in domestic animals and should not be used alone in the assessment of capture myopathy and muscular trauma. Additionally, the dry chemistry analyzer, typically used in clinics, reported significantly higher and less precise AST and ALT activities with poor correlation to those measured with wet chemical methods found in diagnostic laboratories. Therefore, these results cannot be clinically compared. Overall, the optimized wet chemical method was the most precise and diagnostically useful measurement of aminotransferase in samples. Additionally, there was a statistically significant difference between paired serum and plasma measurement

  6. Ultradian rhythmicity of tyrosine aminotransferase activity in Euglena gracillis: Analysis by cosine and non-sinusoidal fitting procedures

    NASA Astrophysics Data System (ADS)

    Neuhaus-Steinmetz, Ulrich; Balzer, Ivonne; Hardeland, Rüdiger

    1990-03-01

    Although the geophysical periodicity of the earth's rotation corresponds to a biological cyclicity of ca. 24 h, cellular temporal organization comprises a multifrequency time structure, in which ultradian rhythms may be regarded as subelements of the circadian oscillator. In Euglena gracilis kept under conditons in which various cellular functions oscillate with a circadian period, tyrosine aminotransferase activity exhibited predominantly an ultradian cycle, whereas its circadian frequency was only weakly expressed. Ultradian period lengths were in the range of 4 5 h, as demonstrated by least squares fitting of cosines and of a non-sinusoidal regression function.

  7. Evaluation of Aspartate Aminotransferase-to-Platelet Ratio Index as a Non-Invasive Marker for Liver Cirrhosis

    PubMed Central

    Tripathi, B.K.; Gupta, B.; Bhandari, Bharti; Jalan, Divesh

    2015-01-01

    Introduction Liver biopsy is considered as a gold standard for the diagnosis of cirrhosis. Till date there is no non-invasive marker to replace it. Aim To investigate the effectiveness of Aspartate aminotransferase-to-platelet ratio index (APRI) as a non-invasive marker for liver cirrhosis. Materials and Methods Fifty-one patients with cirrhosis, identified on USG abdomen were included in study. Platelet count and Aspartate aminotransferase (AST) were done using haematology automatic analyser and automatic HITACHI-912 Auto Analyser respectively. APRI was calculated for every patient using the formula {(AST / ULN) x 100}/platelet count (109/L). Predictive accuracy was evaluated with a receiver-operating characteristics (ROC) curve. Results APRI correctly classified 49 (96.1%) patients of cirrhosis with area under the ROC curve of 0.973 (95% CI) at cut-off 0.65 with negative predictive value (NPV) and Positive predictive value (PPV) of 96% and 96.1% respectively. The sensitivity and specificity of the test was found to be 96% and 96.1% respectively. Conclusion APRI could identify cirrhosis with high degree of accuracy in the studied patients. PMID:26672800

  8. Glucocorticoids locally disrupt an array of positioned nucleosomes on the rat tyrosine aminotransferase promoter in hepatoma cells.

    PubMed Central

    Carr, K D; Richard-Foy, H

    1990-01-01

    Transcriptional activation by steroid hormones is often associated with the appearance of a DNase I hypersensitive site resulting from a local alteration of the nucleoprotein structure of the promoter. For the mouse mammary tumor virus long terminal repeat, a viral promoter under glucocorticoid control, a model has been proposed: the appearance of the hormonodependent DNase I hypersensitive site reflects the displacement of a single precisely positioned nucleosome associated with the glucocorticoid responsive elements. To determine if such a mechanism is of general relevance in transcriptional activation by steroid hormones, we have investigated the nucleosomal organization of the rat tyrosine aminotransferase promoter over a 1-kilobase region that contains the glucocorticoid regulatory target. This region displays a hormonodependent DNase I hypersensitive site. In the absence of hormone, micrococcal nuclease digestion of nuclei demonstrates the presence of positioned nucleosomes, with cutting sites centered around positions -3080, -2900, -2700, -2800, -2255, and -2040. Treatment of the cells with dexamethasone induces a disruption of the chromatin structure over a relatively short stretch of DNA (approximately positions -2400 to -2650) that overlaps two nucleosomes. These observations suggest a strong similarity in the role of chromatin structure in glucocorticoid-dependent transcriptional activation of mouse mammary tumor virus and tyrosine aminotransferase promoters. Images PMID:1979170

  9. Overproduction and preliminary crystallographic study of a human kynurenine aminotransferase II homologue from Pyrococcus horikoshii OT3

    SciTech Connect

    Chon, Hyongi; Matsumura, Hiroyoshi; Shimizu, Shoko; Maeda, Nao; Koga, Yuichi; Takano, Kazufumi; Kanaya, Shigenori

    2005-03-01

    A human kynurenine aminotransferase II homologue from P. horikoshii OT3 has been overproduced in E. coli, purified, and characterized. Crystals of this protein have been obtained and analyzed by X-ray diffraction. The Pyrococcus horikoshii OT3 genome contains a gene encoding a human kynurenine aminotransferase II (KAT II) homologue, which consists of 428 amino-acid residues and shows an amino-acid sequence identity of 30% to human KAT II. This gene was overexpressed in Escherichia coli and the recombinant protein (Ph-KAT II) was purified. Gel-filtration chromatography showed that Ph-KAT II exists as a homodimer. Ph-KAT II exhibited enzymatic activity that catalyzes the transamination of l-kynurenine to produce kynurenic acid. Crystals of Ph-KAT II were grown using the sitting-drop vapour-diffusion method and native X-ray diffraction data were collected to 2.2 Å resolution using synchrotron radiation from station BL44XU at SPring-8. The crystals belong to the centred orthorhombic space group C222{sub 1}, with unit-cell parameters a = 71.75, b = 86.84, c = 137.30 Å. Assuming one molecule per asymmetric unit, the V{sub M} value was 2.19 Å{sup 3} Da{sup −1} and the solvent content was 43.3%.

  10. Structure of ALD1, a plant-specific homologue of the universal diaminopimelate aminotransferase enzyme of lysine biosynthesis.

    PubMed

    Sobolev, Vladimir; Edelman, Marvin; Dym, Orly; Unger, Tamar; Albeck, Shira; Kirma, Menny; Galili, Gad

    2013-02-01

    Diaminopimelate aminotransferase (DAP-AT) is an enzyme in the lysine-biosynthesis pathway. Conversely, ALD1, a close homologue of DAP-AT in plants, uses lysine as a substrate in vitro. Both proteins require pyridoxal-5'-phosphate (PLP) for their activity. The structure of ALD1 from the flowering plant Arabidopsis thaliana (AtALD1) was solved at a resolution of 2.3 Å. Comparison of AtALD1 with the previously solved structure of A. thaliana DAP-AT (AtDAP-AT) revealed similar interactions with PLP despite sequence differences within the PLP-binding site. However, sequence differences between the binding site of AtDAP-AT for malate, a purported mimic of substrate binding, and the corresponding site in AtALD1 led to different interactions. This suggests that either the substrate itself, or the substrate-binding mode, differs in the two proteins, supporting the known in vitro findings.

  11. Mutational analysis of Mycobacterium tuberculosis lysine ɛ-aminotransferase and inhibitor co-crystal structures, reveals distinct binding modes.

    PubMed

    Tripathi, Sarvind Mani; Agarwal, Aparna; Ramachandran, Ravishankar

    Lysine ɛ-aminotransferase (LAT) converts lysine to α-aminoadipate-δ-semialdehyde in a PLP-mediated reaction. We mutated active-site T330, N328 and E243, and structurally rationalized their properties. T330A and T330S mutants cannot bind PLP and are inactive. N328A although inactive, binds to PLP. E243A retains activity, but binds α-ketoglutarate in a different conformation. We had earlier identified 2-aminomethyl piperidine derivative as a LAT inhibitor. The co-crystal structure reveals that it mimics binding of C5 substrates and exhibits two binding modes. E243, that shields R422 in the apo enzyme, exhibits conformational changes to permit the binding of the inhibitor in one of the binding modes. Structure-based analysis of bound water in the active site suggests optimization strategies for synthesis of improved inhibitors.

  12. Cloning Two Genes for Nicotianamine Aminotransferase, a Critical Enzyme in Iron Acquisition (Strategy II) in Graminaceous Plants

    PubMed Central

    Takahashi, Michiko; Yamaguchi, Hirotaka; Nakanishi, Hiromi; Shioiri, Takayuki; Nishizawa, Naoko-Kishi; Mori, Satoshi

    1999-01-01

    Nicotianamine aminotransferase (NAAT), the key enzyme involved in the biosynthesis of mugineic acid family phytosiderophores (MAs), catalyzes the amino transfer of nicotianamine (NA). MAs are found only in graminaceous plants, although NA has been detected in every plant so far investigated. Therefore, this amino transfer reaction is the first step in the unique biosynthesis of MAs that has evolved in graminaceous plants. NAAT activity is dramatically induced by Fe deficiency and suppressed by Fe resupply. Based on the protein sequence of NAAT purified from Fe-deficient barley (Hordeum vulgare) roots, two distinct cDNA clones encoding NAAT, naat-A and naat-B, were identified. Their deduced amino acid sequences were homologous to several aminotransferases, and shared consensus sequences for the pyridoxal phosphate-binding site lysine residue and its surrounding residues. The expression of both naat-A and naat-B is increased in Fe-deficient barley roots, while naat-B has a low level of constitutive expression in Fe-sufficient barley roots. No detectable mRNA from either naat-A or naat-B was present in the leaves of either Fe-deficient or Fe-sufficient barley. One genomic clone with a tandem array of naat-B and naat-A in this order was identified. naat-B and naat-A each have six introns at the same locations. The isolation of NAAT genes will pave the way to understanding the mechanism of the response to Fe in graminaceous plants, and may lead to the development of cultivars tolerant to Fe deficiency that can grow in calcareous soils. PMID:10557244

  13. Ceruloplasmin, a reliable marker of fibrosis in chronic hepatitis B virus patients with normal or minimally raised alanine aminotransferase

    PubMed Central

    Zeng, Da-Wu; Dong, Jing; Jiang, Jia-Ji; Zhu, Yue-Yong; Liu, Yu-Rui

    2016-01-01

    AIM To develop a non-invasive model to evaluate significant fibrosis and cirrhosis by investigating the association between serum ceruloplasmin (CP) levels and liver fibrosis in chronic hepatitis B (CHB) patients with normal or minimally raised alanine aminotransferase (ALT). METHODS Serum samples and liver biopsy were obtained from 193 CHB patients with minimally raised or normal ALT who were randomly divided into a training group (n = 97) and a validation group (n = 96). Liver histology was evaluated by the METAVIR scoring system. Receiver operator characteristic curves were applied to the diagnostic value of CP for measuring liver fibrosis in CHB patients. Spearman rank correlation analyzed the relationship between CP and liver fibrosis. A non-invasive model was set up through multivariate logistic regression analysis. RESULTS Serum CP levels individualized various fibrosis stages via area under the curve (AUC) values. Multivariate analysis revealed that CP levels were significantly related to liver cirrhosis. Combining CP with serum GGT levels, a CG model was set up to predict significant fibrosis and liver cirrhosis in CHB patients with normal or minimally raised ALT. The AUC, sensitivity, specificity, positive predictive value, and negative predictive value were 0.84, 83.1%, 78.6%, 39.6%, and 96.5% to predict liver cirrhosis, and 0.789, 80.26%, 68.38%, 62.25%, and 84.21% to predict significant fibrosis. This model expressed a higher AUC than FIB-4 (age, ALT, aspartate aminotransferase, platelets) and GP (globulin, platelets) models to predict significant fibrosis (P = 0.019 and 0.022 respectively) and revealed a dramatically greater AUC than FIB-4 (P = 0.033) to predict liver cirrhosis. CONCLUSION The present study showed that CP was independently and negatively associated with liver fibrosis. Furthermore, we developed a novel promising model (CG), based on routine serum markers, for predicting liver fibrosis in CHB patients with normal or minimally raised

  14. Taurine-like GABA aminotransferase inhibitors prevent rabbit brain slices against oxygen-glucose deprivation-induced damage.

    PubMed

    Ricci, Lorenzo; Valoti, Massimo; Sgaragli, Giampietro; Frosini, Maria

    2012-06-01

    The activation of the GABAergic system has been shown to protect brain tissues against the damage that occurs after cerebral ischaemia. On the other hand, the taurine analogues (±)Piperidine-3-sulphonic- (PSA), 2-aminoethane phosphonic- (AEP), 2-(N-acetylamino) cyclohexane sulfonic-acids (ATAHS) and 2-aminobenzene sulfonate-acids (ANSA) have been reported to block GABA metabolism by inhibiting rabbit brain GABA aminotransferase and to increase GABA content in rabbit brain slices. The present investigation explored the neuroprotection provided by GABA, Vigabatrin (VIGA) and taurine analogues in the course of oxygen-glucose deprivation and reperfusion induced damage of rabbit brain slices. Tissue damage was assessed by measuring the release of glutamate and lactate dehydrogenase (LDH) during reperfusion and by determining final tissue water gain, measured as the index of cell swelling. GABA (30-300 μM) and VIGA (30-300 μM) significantly antagonised LDH and glutamate release, as well as tissue water gain caused by oxygen-glucose deprivation and reperfusion. Lower (1-10 μM) or higher concentrations (up to 3,000 μM) were ineffective. ANSA, PSA and ATAHS significantly reduced glutamate and LDH release and tissue water gain in a range of concentrations between 30 and 300 μM. Lower (0-10 μM) or higher (up to 3,000 μM) concentrations were ineffective. Both mechanisms suggest hormetic ("U-shaped") effects. These results indicate that the GABAergic system activation performed directly by GABA or indirectly through GABA aminotransferase inhibition is a promising approach for protecting the brain against ischemia and reperfusion-induced damage.

  15. Prevalence and Predictors of Elevated Aspartate Aminotransferase-to-Platelet Ratio Index in Latin American Perinatally HIV-infected Children

    PubMed Central

    Siberry, George K.; Cohen, Rachel A.; Harris, D. Robert; Cruz, Maria Leticia Santos; Oliveira, Ricardo; Peixoto, Mario F.; Cervi, Maria Celia; Hazra, Rohan; Pinto, Jorge A.

    2013-01-01

    Background Chronic liver disease has emerged as an important problem in adults with longstanding HIV infection, but data are lacking for children. We characterized elevated aspartate aminotransferase (AST)-to-platelet ratio index (APRI ), a marker of possible liver fibrosis, in perinatally HIV-infected children. Methods NISDI [NICHD (National Institute of Child Health and Human Development) International Site Development Initiative] enrolled HIV-infected children (ages 0.1-20.1 years) from five Latin American countries in an observational cohort from 2002–2009. Twice yearly visits included medical history, physical examination and laboratory evaluations. The prevalence (95% confidence interval [CI]) of APRI>1.5 was calculated and associations with demographic, HIV-related and liver-related variables were investigated in bivariate analyses. Results APRI was available for 1012 of 1032 children. APRI was >1.5 in 32 (3.2%, 95% CI: 2.2%-4.4%) including 2 of 4 participants with hepatitis B (HBV) infection. Factors significantly associated with APRI>1.5 (p<0.01 compared to APRI≤1.5) included country, younger age, past or current HBV, higher alanine aminotransferase, lower total cholesterol, higher log10 current viral load, lower current CD4 count, lower nadir CD4 count, use of hepatotoxic non-antiretroviral (ARV) medications, and no prior ARV use. Rates of APRI>1.5 varied significantly by current ARV regimen (p=0.0002), from 8.0% for no ARV to 3.2% for non-protease inhibitor (PI) regimens to 1.5% for PI-based regimens. Conclusions Elevated APRI occurred in approximately 3% of perinatally HIV-infected children. PI-based ARVs appeared protective while inadequate HIV control appeared to increase risk of elevated APRI. Additional investigations are needed to better assess potential subclinical, chronic liver disease in HIV-infected children. PMID:23799515

  16. Assignment of the rat genes coding for phenylalanine hydroxylase (PAH), tyrosine aminotransferase (TAT), and pyruvate kinase (PKL) to chromosomes 7, 19, 2, respectively.

    PubMed

    Fulchignoni-Lataud, M C; Weiss, M C; Szpirer, C; Levan, G

    1990-01-01

    A panel of hybrid clones segregating rat chromosomes in a mouse background was used to determine the chromosomal localization of three genes specifically expressed in hepatocytes. The phenylalanine hydroxylase, tyrosine aminotransferase, and pyruvate kinase genes were assigned to rat chromosomes 7, 19, and 2, respectively.

  17. Comparison of effect of cafetière and filtered coffee on serum concentrations of liver aminotransferases and lipids: six month randomised controlled trial.

    PubMed Central

    Urgert, R.; Meyboom, S.; Kuilman, M.; Rexwinkel, H.; Vissers, M. N.; Klerk, M.; Katan, M. B.

    1996-01-01

    OBJECTIVE: To study the effects of prolonged intake of cafetière coffee, which is rich in the diterpenes cafestol and kahweol, on serum aminotransferase and lipid concentrations. DESIGN: Randomised parallel controlled trial. SUBJECTS: 46 healthy men and women aged 19 to 69. INTERVENTION: Consumption of five to six strong cups (0.9 litres) a day of either cafetière (22 subjects) or filtered coffee (24 subjects) for 24 weeks. MAIN OUTCOME MEASURES: Mean changes in serum aminotransferase and lipid concentrations. RESULTS: Cafetière coffee raised alanine aminotransferase concentration by up to 80% above baseline values relative to filtered coffee. After 24 weeks the rise was still 45% (9 U/l (95% confidence interval 3 to 15 U/l), P = 0.007). Alanine aminotransferase concentration exceeded the upper limit of normal in eight of the 22 subjects drinking cafetière coffee, being twice the upper limit of normal in three of them. Cafetière coffee raised low density lipoprotein cholesterol concentrations by 9-14%. After 24 weeks the rise was 0.26 mmol/l (0.04 to 0.47 mmol/l) (P = 0.03) relative to filtered coffee. Triglyceride concentrations initially rose by 26% with cafetière coffee but returned close to baseline values within six months. All increases were reversible after the intervention was stopped. CONCLUSIONS: Daily consumption of five to six cups of strong cafetière coffee affects the integrity of liver cells as suggested by small increases in serum alanine aminotransferase concentration. The effect does not subside with prolonged intake. High intakes of coffee brews rich in cafestol and kahweol may thus be responsible for unexplained increases in this enzyme activity in apparently healthy subjects. Cafetière coffee also raises low density lipoprotein cholesterol concentration and thus the risk of coronary heart disease. PMID:8956701

  18. Preoperative aspartate aminotransferase-to-platelet ratio index (APRI) is a predictor on postoperative outcomes of hepatocellular carcinoma

    PubMed Central

    Cheng, JiWen; Zhao, Pu; Liu, JiangBo; Liu, Xi; Wu, XuanLin

    2016-01-01

    Abstract Preoperative aspartate aminotransferase-to-platelet ratio index (APRI) has been identified as a biochemical marker for histological fibrogenesis and fibrosis in cirrhosis and prognosis of hepatocellular carcinoma (HCC). Whether preoperative APRI can predict postoperative short-term outcomes has not been studied. The purpose of this study was to investigate the ability of preoperative APRI to predict short-term outcomes following liver resection for HCC. APRI was evaluated in 360 patients undergoing liver resection for HCC. The receiver operating characteristic curve analysis was conducted to determine the cutoff value of the APRI in predicting postoperative morbidity. Univariate and multivariate analysis was performed to identify the risk factors for postoperative outcomes. The correlation of the preoperative APRI value with clinicopathological parameters was also examined. We found that the optimal cutoff value of the APRI was set at 9.5 for postoperative complications. APRI was an independent risk factor for overall complications by univariate and multivariate analyses. HCC patients with elevated APRI (>9.5) had a worse liver function and significantly higher postoperative complication rate. In conclusion, preoperative APRI is a useful biochemical marker to predict postoperative outcomes in HCC patients. PMID:27902606

  19. Crystallization and preliminary X-ray diffraction analysis of l,l-diaminopimelate aminotransferase (DapL) from Chlamydomonas reinhardtii

    PubMed Central

    Hudson, André O.; Girón, Irma; Dobson, Renwick C. J.

    2011-01-01

    In the anabolic synthesis of diaminopimelate and lysine in plants and in some bacteria, the enzyme l,l-diaminopimelate aminotransferase (DapL; EC 2.6.1.83) catalyzes the conversion of tetrahydrodipicolinic acid (THDPA) to l,l-diaminopimelate, bypassing the DapD, DapC and DapE enzymatic steps in the bacterial acyl pathways. Here, the cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of DapL from the alga Chlamydomonas reinhardtii are presented. Protein crystals were grown in conditions containing 25%(w/v) PEG 3350 and 200 mM lithium sulfate and initially diffracted to ∼1.35 Å resolution. They belonged to space group P212121, with unit-cell parameters a = 58.9, b = 91.8, c = 162.9 Å. The data were processed to 1.55 Å resolution with an R merge of 0.081, an R p.i.m. of 0.044, an R r.i.m of 0.093 and a V M of 2.28 Å3 Da−1. PMID:21206046

  20. Crystallization and preliminary X-ray diffraction analysis of L,L-diaminopimelate aminotransferase (DapL) from Chlamydomonas reinhardtii.

    PubMed

    Hudson, André O; Girón, Irma; Dobson, Renwick C J

    2011-01-01

    In the anabolic synthesis of diaminopimelate and lysine in plants and in some bacteria, the enzyme L,L-diaminopimelate aminotransferase (DapL; EC 2.6.1.83) catalyzes the conversion of tetrahydrodipicolinic acid (THDPA) to L,L-diaminopimelate, bypassing the DapD, DapC and DapE enzymatic steps in the bacterial acyl pathways. Here, the cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of DapL from the alga Chlamydomonas reinhardtii are presented. Protein crystals were grown in conditions containing 25% (w/v) PEG 3350 and 200 mM lithium sulfate and initially diffracted to ∼1.35 Å resolution. They belonged to space group P2(1)2(1)2(1), with unit-cell parameters a=58.9, b=91.8, c=162.9 Å. The data were processed to 1.55 Å resolution with an Rmerge of 0.081, an Rp.i.m. of 0.044, an Rr.i.m of 0.093 and a VM of 2.28 Å3 Da(-1).

  1. Genetic engineering of improved nitrogen use efficiency in rice by the tissue-specific expression of alanine aminotransferase.

    PubMed

    Shrawat, Ashok K; Carroll, Rebecka T; DePauw, Mary; Taylor, Gregory J; Good, Allen G

    2008-09-01

    Summary Nitrogen is quantitatively the most essential nutrient for plants and a major factor limiting crop productivity. One of the critical steps limiting the efficient use of nitrogen is the ability of plants to acquire it from applied fertilizer. Therefore, the development of crop plants that absorb and use nitrogen more efficiently has been a long-term goal of agricultural research. In an attempt to develop nitrogen-efficient plants, rice (Oryza sativa L.) was genetically engineered by introducing a barley AlaAT (alanine aminotransferase) cDNA driven by a rice tissue-specific promoter (OsAnt1). This modification increased the biomass and grain yield significantly in comparison with control plants when plants were well supplied with nitrogen. Compared with controls, transgenic rice plants also demonstrated significant changes in key metabolites and total nitrogen content, indicating increased nitrogen uptake efficiency. The development of crop plants that take up and assimilate nitrogen more efficiently would not only improve the use of nitrogen fertilizers, resulting in lower production costs, but would also have significant environmental benefits. These results are discussed in terms of their relevance to the development of strategies to engineer enhanced nitrogen use efficiency in crop plants.

  2. In situ detection of myocardial infarction in pig by measurements of aspartate aminotransferase (ASAT) activity in the interstitial fluid.

    PubMed

    Kennergren, C; Nyström, B; Nyström, U; Berglin, E; Larsson, G; Mantovani, V; Lönnroth, P; Hamberger, A

    1997-01-01

    Microdialysis probes permeable to large molecules (m.w. cut-off > 200 kD) were introduced into the myocardium of anaesthetized pigs in order to evaluate their potential for early detection of myocardial ischaemia and enzyme markers for infarction. The left anterior descending coronary artery was occluded for 30 min and the myocardium was reperfused for 3 h. The concentrations of aspartate aminotransferase (ASAT), lactate, glucose and selected free amino acids were measured. The levels in the interstitium of ischaemic and non-ischaemic myocardium were compared with those in plasma from the coronary sinus as well as from a peripheral vein. Twelve probes were inserted in six pigs and withdrawn after 8-72 hours of sampling. No complications occurred. Simultaneous 100% increase of ASAT and lactate was found in myocardial dialysates after 30 min of ischaemia. ASAT activity remained at that level until the end of reperfusion. The plasma peak ASAT level was not attained until after 3 h. Glutamate was the only amino acid which increased significantly in the myocardial interstitium during ischaemia, peaking after 30 min of reperfusion. Dialysates from the unaffected myocardium showed no effects on lactate, ASAT or glutamate. The use of myocardial microdialysis for pre- and postoperative recordings in man is discussed.

  3. Liver peroxisomal alanine:glyoxylate aminotransferase and the effects of mutations associated with Primary Hyperoxaluria Type I: An overview.

    PubMed

    Oppici, Elisa; Montioli, Riccardo; Cellini, Barbara

    2015-09-01

    Liver peroxisomal alanine:glyoxylate aminotransferase (AGT) (EC 2.6.1.44) catalyses the conversion of l-alanine and glyoxylate to pyruvate and glycine, a reaction that allows glyoxylate detoxification. Inherited mutations on the AGXT gene encoding AGT lead to Primary Hyperoxaluria Type I (PH1), a rare disorder characterized by the deposition of calcium oxalate crystals primarily in the urinary tract. Here we describe the results obtained on the biochemical features of AGT as well as on the molecular and cellular effects of polymorphic and pathogenic mutations. A complex scenario on the molecular pathogenesis of PH1 emerges in which the co-inheritance of polymorphic changes and the condition of homozygosis or compound heterozygosis are two important factors that determine the enzymatic phenotype of PH1 patients. All the reported data represent relevant steps toward the understanding of genotype/phenotype correlations, the prediction of the response of the patients to the available therapies, and the development of new therapeutic approaches. This article is part of a Special Issue entitled: Cofactor-dependent proteins: evolution, chemical diversity and bio-applications.

  4. Distribution of messenger RNAs encoding the enzymes glutaminase, aspartate aminotransferase and glutamic acid decarboxylase in rat brain.

    PubMed

    Najlerahim, A; Harrison, P J; Barton, A J; Heffernan, J; Pearson, R C

    1990-05-01

    In situ hybridization histochemistry (ISHH) using synthetic oligonucleotide probes has been used to identify cells containing the mRNAs coding for glutaminase (GluT), aspartate aminotransferase (AspT) and glutamic acid decarboxylase (GAD). The distribution of GAD mRNA confirms previous descriptions and matches the distribution of GAD detected using specific antibodies. AspT mRNA is widely distributed in the brain, but is present at high levels in GABAergic neuronal populations, some that may be glutamatergic, and in a subset of neurons which do not contain significant levels of either GAD or GluT mRNA. Particularly prominent are the neurons of the magnocellular division of the red nucleus, the large cells in the deep cerebellar nuclei and the vestibular nuclei and neurons of the lateral superior olivary nucleus. GluT mRNA does not appear to be present at high levels in all GAD-containing neurons, but is seen prominently in many neuronal populations that may use glutamate as a neurotransmitter, such as neocortical and hippocampal pyramidal cells, the granule cells of the cerebellum and neurons of the dentate gyrus of the hippocampus. The heaviest labelling of GluT mRNA is seen in the lateral reticular nucleus of the medulla. ISHH using probes directed against the mRNAs encoding these enzymes may be an important technique for identifying glutamate and aspartate using neuronal populations and for examining their regulation in a variety of experimental and pathological circumstances.

  5. The aspartate aminotransferase family in conifers: biochemical analysis of a prokaryotic-type enzyme from maritime pine.

    PubMed

    de la Torre, Fernando; Suárez, María Fernanda; Santis, Laura de; Cánovas, Francisco M

    2007-09-01

    Plant aspartate aminotransferase (AAT, EC 2.6.1.1) plays a key role in primary nitrogen assimilation, the transfer of reducing equivalents and the interchanges of carbon and nitrogen pools between subcellular compartments. We investigated the AAT family in conifers using maritime pine as the experimental model. Genes for cytosolic, mitochondrial and two plastidic isoenzymes (eukaryotic- and prokaryotic-types) were identified and their deduced amino acid sequences compared. The primary structure of the eukaryotic-type enzymes is quite well conserved, whereas the prokaryotic-type AAT is highly divergent (15% of identity). These molecular data were confirmed by the absence of immunological cross-reactivity between the two types of native AATs. The mature prokaryotic-type polypeptide was overexpressed in Escherichia coli, and the native enzyme was purified to apparent homogeneity and its molecular properties determined. The fully active recombinant holoenzyme showed highest catalytic activity at 50-60 degrees C and was moderately thermostable, retaining about 50% of its activity after incubation at 70 degrees C for 5-10 min. The presence of pyridoxal 5'-phosphate significantly increased the thermostability of the enzyme. These molecular characteristics were exploited to develop a rapid protocol for the purification of this prokaryotic-type enzyme from pine cotyledons. The results will be useful for studying aspartate and amino acid metabolism in trees.

  6. Identification and functional analysis of a prokaryotic-type aspartate aminotransferase: implications for plant amino acid metabolism.

    PubMed

    de la Torre, Fernando; De Santis, Laura; Suárez, María Fernanda; Crespillo, Remedios; Cánovas, Francisco M

    2006-05-01

    In this paper, we report the identification of genes from pine (PpAAT), Arabidopsis (AtAAT) and rice (OsAAT) encoding a novel class of aspartate aminotransferase (AAT, EC 2.6.1.1) in plants. The enzyme is unrelated to other eukaryotic AATs from plants and animals but similar to bacterial enzymes. Phylogenetic analysis indicates that this prokaryotic-type AAT is closely related to cyanobacterial enzymes, suggesting it might have an endosymbiotic origin. Interestingly, most of the essential residues involved in the interaction with the substrate and the attachment of pyridoxal phosphate cofactor in the active site of the enzyme were conserved in the deduced polypeptide. The polypeptide is processed in planta to a mature subunit of 45 kDa that is immunologically distinct from the cytosolic, mitochondrial and chloroplastic isoforms of AAT previously characterized in plants. Functional expression of PpAAT sequences in Escherichia coli showed that the processed precursor is assembled into a catalytically active homodimeric holoenzyme that is strictly specific for aspartate. These atypical genes are predominantly expressed in green tissues of pine, Arabidopsis and rice, suggesting a key role of this AAT in nitrogen metabolism associated with photosynthetic activity. Moreover, immunological analyses revealed that the plant prokaryotic-type AAT is a nuclear-encoded chloroplast protein. This implies that two plastidic AAT co-exist in plants: a eukaryotic type previously characterized and the prokaryotic type described here. The respective roles of these two enzymes in plant amino acid metabolism are discussed.

  7. Relationship of creatine kinase, aspartate aminotransferase, lactate dehydrogenase, and proteinuria to cardiomyopathy in the owl monkey (Aotus vociferans)

    SciTech Connect

    Gozalo, Alfonso S.; Chavera, Alfonso; Montoya, Enrique J.; Takano, Juan; Weller, Richard E.

    2008-02-01

    The purpose of this study was to determine serum reference values for crea- tine kinase (CK), aspartate aminotransferase (AST), and lactate dehydroge- nase (LDH) in captive-born and wild-caught owl monkeys to assess their usefulness for diagnosing myocardial disease. Urine samples were also collected and semi-quantitative tests performed. There was no statistically significant difference between CK, AST, and LDH when comparing both groups. However, when comparing monkeys with proteinuria to those without proteinuria, a statistically significant difference in CK value was observed (P = 0.021). In addition, the CK/AST ratio revealed that 29% of the animals included in this study had values suggesting cardiac infarction. Grossly, cardiac concentric hypertrophy of the left ventricle and small, pitted kidneys were the most common findings. Microscopically, myocardial fibrosis, contraction band necrosis, hypertrophy and hyperplasia of coronary arteries, medium-sized renal arteries, and afferent glomerular arteriolae were the most significant lesions, along with increased mesangial matrix and hypercellularity of glomeruli, Bowman’s capsule, and peritubular space fibroplasia. These findings suggest that CK, AST, and LDH along with urinalysis provide a reliable method for diagnosing cardiomyopathies in the owl monkey. In addition, CK/AST ratio, proteinuria, and the observed histological and ultrastructural changes suggest that Aotus vociferans suffer from arterial hypertension and chronic myocardial infarction.

  8. A root-expressed L-phenylalanine:4-hydroxyphenylpyruvate aminotransferase is required for tropane alkaloid biosynthesis in Atropa belladonna.

    PubMed

    Bedewitz, Matthew A; Góngora-Castillo, Elsa; Uebler, Joseph B; Gonzales-Vigil, Eliana; Wiegert-Rininger, Krystle E; Childs, Kevin L; Hamilton, John P; Vaillancourt, Brieanne; Yeo, Yun-Soo; Chappell, Joseph; DellaPenna, Dean; Jones, A Daniel; Buell, C Robin; Barry, Cornelius S

    2014-09-01

    The tropane alkaloids, hyoscyamine and scopolamine, are medicinal compounds that are the active components of several therapeutics. Hyoscyamine and scopolamine are synthesized in the roots of specific genera of the Solanaceae in a multistep pathway that is only partially elucidated. To facilitate greater understanding of tropane alkaloid biosynthesis, a de novo transcriptome assembly was developed for Deadly Nightshade (Atropa belladonna). Littorine is a key intermediate in hyoscyamine and scopolamine biosynthesis that is produced by the condensation of tropine and phenyllactic acid. Phenyllactic acid is derived from phenylalanine via its transamination to phenylpyruvate, and mining of the transcriptome identified a phylogenetically distinct aromatic amino acid aminotransferase (ArAT), designated Ab-ArAT4, that is coexpressed with known tropane alkaloid biosynthesis genes in the roots of A. belladonna. Silencing of Ab-ArAT4 disrupted synthesis of hyoscyamine and scopolamine through reduction of phenyllactic acid levels. Recombinant Ab-ArAT4 preferentially catalyzes the first step in phenyllactic acid synthesis, the transamination of phenylalanine to phenylpyruvate. However, rather than utilizing the typical keto-acid cosubstrates, 2-oxoglutarate, pyruvate, and oxaloacetate, Ab-ArAT4 possesses strong substrate preference and highest activity with the aromatic keto-acid, 4-hydroxyphenylpyruvate. Thus, Ab-ArAT4 operates at the interface between primary and specialized metabolism, contributing to both tropane alkaloid biosynthesis and the direct conversion of phenylalanine to tyrosine.

  9. The tryptophan aminotransferase Tam1 catalyses the single biosynthetic step for tryptophan-dependent pigment synthesis in Ustilago maydis.

    PubMed

    Zuther, Katja; Mayser, Peter; Hettwer, Ursula; Wu, Wenying; Spiteller, Peter; Kindler, Bernhard L J; Karlovsky, Petr; Basse, Christoph W; Schirawski, Jan

    2008-04-01

    Tryptophan is a precursor for many biologically active secondary metabolites. We have investigated the origin of indole pigments first described in the pityriasis versicolor-associated fungus Malassezia furfur. Some of the identified indole pigments have properties potentially explaining characteristics of the disease. As M. furfur is not amenable to genetic manipulation, we used Ustilago maydis to investigate the pathway leading to pigment production from tryptophan. We show by high-performance liquid chromatography, mass spectrometry and nuclear magnetic resonance analysis that the compounds produced by U. maydis include those putatively involved in the etiology of pityriasis versicolor. Using a reverse genetics approach, we demonstrate that the tryptophan aminotransferase Tam1 catalyses pigment biosynthesis by conversion of tryptophan into indolepyruvate. A forward genetics approach led to the identification of mutants incapable of producing the pigments. These mutants were affected in the sir1 gene, presumably encoding a sulphite reductase. In vitro experiments with purified Tam1 showed that 2-oxo 4-methylthio butanoate serves as a substrate linking tryptophan deamination to sulphur metabolism. We provide the first direct evidence that these indole pigments form spontaneously from indolepyruvate and tryptophan without any enzymatic activity. This suggests that compounds with a proposed function in M. furfur-associated disease consist of indolepyruvate-derived spontaneously generated metabolic by-products.

  10. Ammonia upregulates kynurenine aminotransferase II mRNA expression in rat brain: a role for astrocytic NMDA receptors?

    PubMed

    Obara-Michlewska, Marta; Tuszyńska, Paulina; Albrecht, Jan

    2013-06-01

    Kynurenine aminotransferase II (KAT-II) is the astrocytic enzyme catalyzing the synthesis of kynurenic acid (KYNA), an endogenous inhibitor of the α7-nicotinic receptor and the NMDA receptor (NMDAr). A previous study demonstrated an increase of KYNA synthesis in the brain of rats with thioacetamide (TAA)-induced acute liver failure. Here we show that TAA administration increases KAT-II expression in the rat cerebral cortex and the effect is mimicked in cerebral cortical astrocytes in culture treated with high (5 mM) concentration of ammonia. KAT-II expression in control and TAA-treated rats was increased by NMDAr antagonist memantine, and the effects of TAA and memantine appeared additive. In astrocytes, the NMDAr antagonist MK-801 raised KAT-II expression as well, while NMDA added alone had no effect. Glutamate decreased KAT-II mRNA level, which was attenuated by MK-801. The results suggest that stimulation of KAT-II expression during hepatic encephalopathy may be associated with a partial inactivation of astrocytic NMDAr by ammonia.

  11. Branched-chain amino acid metabolon: interaction of glutamate dehydrogenase with the mitochondrial branched-chain aminotransferase (BCATm).

    PubMed

    Islam, Mohammad Mainul; Nautiyal, Manisha; Wynn, R Max; Mobley, James A; Chuang, David T; Hutson, Susan M

    2010-01-01

    The catabolic pathway for branched-chain amino acids includes deamination followed by oxidative decarboxylation of the deaminated product branched-chain alpha-keto acids, catalyzed by the mitochondrial branched-chain aminotransferase (BCATm) and branched-chain alpha-keto acid dehydrogenase enzyme complex (BCKDC). We found that BCATm binds to the E1 decarboxylase of BCKDC, forming a metabolon that allows channeling of branched-chain alpha-keto acids from BCATm to E1. The protein complex also contains glutamate dehydrogenase (GDH1), 4-nitrophenylphosphatase domain and non-neuronal SNAP25-like protein homolog 1, pyruvate carboxylase, and BCKDC kinase. GDH1 binds to the pyridoxamine 5'-phosphate (PMP) form of BCATm (PMP-BCATm) but not to the pyridoxal 5'-phosphate-BCATm and other metabolon proteins. Leucine activates GDH1, and oxidative deamination of glutamate is increased further by addition of PMP-BCATm. Isoleucine and valine are not allosteric activators of GDH1, but in the presence of 5'-phosphate-BCATm, they convert BCATm to PMP-BCATm, stimulating GDH1 activity. Sensitivity to ADP activation of GDH1 was unaffected by PMP-BCATm; however, addition of a 3 or higher molar ratio of PMP-BCATm to GDH1 protected GDH1 from GTP inhibition by 50%. Kinetic results suggest that GDH1 facilitates regeneration of the form of BCATm that binds to E1 decarboxylase of the BCKDC, promotes metabolon formation, branched-chain amino acid oxidation, and cycling of nitrogen through glutamate.

  12. A Novel Pathway for Metabolism of the Cardiovascular Risk Factor Homoarginine by alanine:glyoxylate aminotransferase 2

    PubMed Central

    Rodionov, Roman N.; Oppici, Elisa; Martens-Lobenhoffer, Jens; Jarzebska, Natalia; Brilloff, Silke; Burdin, Dmitrii; Demyanov, Anton; Kolouschek, Anne; Leiper, James; Maas, Renke; Cellini, Barbara; Weiss, Norbert; Bode-Böger, Stefanie M.

    2016-01-01

    Low plasma concentrations of L-homoarginine are associated with an increased risk of cardiovascular events, while homoarginine supplementation is protective in animal models of metabolic syndrome and stroke. Catabolism of homoarginine is still poorly understood. Based on the recent findings from a Genome Wide Association Study we hypothesized that homoarginine can be metabolized by alanine:glyoxylate aminotransferase 2 (AGXT2). We purified human AGXT2 from tissues of AGXT2 transgenic mice and demonstrated its ability to metabolize homoarginine to 6-guanidino-2-oxocaproic acid (GOCA). After incubation of HepG2 cells overexpressing AGXT2 with isotope-labeled homoarginine-d4 we were able to detect labeled GOCA in the medium. We injected wild type mice with labeled homoarginine and detected labeled GOCA in the plasma. We found that AGXT2 knockout (KO) mice have higher homoarginine and lower GOCA plasma levels as compared to wild type mice, while the reverse was true for AGXT2 transgenic (Tg) mice. In summary, we experimentally proved the presence of a new pathway of homoarginine catabolism – its transamination by AGXT2 with formation of GOCA and demonstrated that endogenous AGXT2 is required for maintenance of homoarginine levels in mice. Our findings may lead to development of novel therapeutic approaches for cardiovascular pathologies associated with homoarginine deficiency. PMID:27752063

  13. Lack of association between TTV viral load and aminotransferase levels in patients with hepatitis C or non-B-C.

    PubMed

    Kato, H; Mizokami, M; Orito, E; Ohno, T; Hayashi, K; Nakano, T; Kato, T; Tanaka, Y; Sugauchi, F; Mukaide, M; Ueda, R

    2000-01-01

    TT virus (TTV) is a newly identified un-enveloped single-stranded DNA virus. Although TTV was initially thought to be a new hepatitis virus, it is still unclear whether it causes hepatitis. To clarify the natural history and pathogenesis of TTV infection, serial serum samples from patients with chronic hepatitis were analysed. TTV DNA was quantified by real-time detection polymerase chain reaction assay (RTD-PCR), which was adapted for TTV. Five patients with chronic hepatitis, 4 with hepatitis C and 1 with non-B-C, were studied. The study period ranged from 9 to 50 months. In 3 patients there were frequent increases in TTV DNA titres, but no concomitant elevation of the aminotransferase (ALT) levels. In 2 patients who were treated with interferon, the changes in TTV titres were not synchronized with those of the ALT levels. Thus, in cases of chronic hepatitis, no correlation was observed between the serum TTV DNA titres and the ALT levels.

  14. Aspartic acid aminotransferase activity is increased in actively spiking compared with non-spiking human epileptic cortex.

    PubMed Central

    Kish, S J; Dixon, L M; Sherwin, A L

    1988-01-01

    Increased concentration of the excitatory neurotransmitter aspartic acid in actively spiking human epileptic cerebral cortex was recently described. In order to further characterise changes in the aspartergic system in epileptic brain, the behaviour of aspartic acid aminotransferase (AAT), a key enzyme involved in aspartic acid metabolism has now been examined. Electrocorticography performed during surgery was employed to identify cortical epileptic spike foci in 16 patients undergoing temporal lobectomy for intractable seizures. Patients with spontaneously spiking lateral temporal cortex (n = 8) were compared with a non-spiking control group (n = 8) of patients in whom the epileptic lesions were confined to the hippocampus sparing the temporal convexity. Mean activity of AAT in spiking cortex was significantly elevated by 16-18%, with aspartic acid concentration increased by 28%. Possible explanations for the enhanced AAT activity include increased proliferation of cortical AAT-containing astrocytes at the spiking focus and/or a generalised increase in neuronal or extraneuronal metabolism consequent to the ongoing epileptic discharge. It is suggested that the data provide additional support for a disturbance of central excitatory aspartic acid mechanisms in human epileptic brain. PMID:2898010

  15. Dietary freshwater clam (Corbicula fluminea) extract suppresses accumulation of hepatic lipids and increases in serum cholesterol and aminotransferase activities induced by dietary chloretone in rats.

    PubMed

    Chijimatsu, Takeshi; Umeki, Miki; Kobayashi, Satoru; Kataoka, Yutaro; Yamada, Koji; Oda, Hiroaki; Mochizuki, Satoshi

    2015-01-01

    We investigated the ameliorative effect of freshwater clam extract (FCE) on fatty liver, hypercholesterolemia, and liver injury in rats exposed to chloretone. Furthermore, we examined the effects of major FCE components (fat and protein fractions) to determine the active components in FCE. Chloretone increased serum aminotransferase activities and led to hepatic lipid accumulation. Serum aminotransferase activities and hepatic lipid content were lower in rats fed total FCE or fat/protein fractions of FCE. Expression of fatty acid synthase and fatty acid desaturase genes was upregulated by chloretone. Total FCE and fat/protein fractions of FCE suppressed the increase in gene expression involved in fatty acid synthesis. Serum cholesterol levels increased twofold upon chloretone exposure. Total FCE or fat/protein fractions of FCE showed hypocholesterolemic effects in rats with hypercholesterolemia induced by chloretone. These suggest that FCE contains at least two active components against fatty liver, hypercholesterolemia, and liver injury in rats exposed to chloretone.

  16. Kynurenine Aminotransferase III and Glutamine Transaminase L Are Identical Enzymes that have Cysteine S-Conjugate β-Lyase Activity and Can Transaminate l-Selenomethionine*

    PubMed Central

    Pinto, John T.; Krasnikov, Boris F.; Alcutt, Steven; Jones, Melanie E.; Dorai, Thambi; Villar, Maria T.; Artigues, Antonio; Li, Jianyong; Cooper, Arthur J. L.

    2014-01-01

    Three of the four kynurenine aminotransferases (KAT I, II, and IV) that synthesize kynurenic acid, a neuromodulator, are identical to glutamine transaminase K (GTK), α-aminoadipate aminotransferase, and mitochondrial aspartate aminotransferase, respectively. GTK/KAT I and aspartate aminotransferase/KAT IV possess cysteine S-conjugate β-lyase activity. The gene for the former enzyme, GTK/KAT I, is listed in mammalian genome data banks as CCBL1 (cysteine conjugate beta-lyase 1). Also listed, despite the fact that no β-lyase activity has been assigned to the encoded protein in the genome data bank, is a CCBL2 (synonym KAT III). We show that human KAT III/CCBL2 possesses cysteine S-conjugate β-lyase activity, as does mouse KAT II. Thus, depending on the nature of the substrate, all four KATs possess cysteine S-conjugate β-lyase activity. These present studies show that KAT III and glutamine transaminase L are identical enzymes. This report also shows that KAT I, II, and III differ in their ability to transaminate methyl-l-selenocysteine (MSC) and l-selenomethionine (SM) to β-methylselenopyruvate (MSP) and α-ketomethylselenobutyrate, respectively. Previous studies have identified these seleno-α-keto acids as potent histone deacetylase inhibitors. Methylselenol (CH3SeH), also purported to have chemopreventive properties, is the γ-elimination product of SM and the β-elimination product of MSC catalyzed by cystathionine γ-lyase (γ-cystathionase). KAT I, II, and III, in part, can catalyze β-elimination reactions with MSC generating CH3SeH. Thus, the anticancer efficacy of MSC and SM will depend, in part, on the endogenous expression of various KAT enzymes and cystathionine γ-lyase present in target tissue coupled with the ability of cells to synthesize in situ either CH3SeH and/or seleno-keto acid metabolites. PMID:25231977

  17. Cloning, purification, crystallization and preliminary X-ray crystallographic analysis of the biosynthetic N-acetylornithine aminotransferases from Salmonella typhimurium and Escherichia coli

    SciTech Connect

    Rajaram, V.; Prasad, K.; Ratna Prasuna, P.; Ramachandra, N.; Bharath, S. R.; Savithri, H. S.; Murthy, M. R. N.

    2006-10-01

    Acetylornithine aminotransferases, members of the type I subgroup II family of PLP-dependent enzymes, from S. typhimurium and E. coli have been cloned, overexpressed, purified and crystallized. Acetylornithine aminotransferase (AcOAT) is a type I pyridoxal 5′-phosphate-dependent enzyme catalyzing the conversion of N-acetylglutamic semialdehyde to N-acetylornithine in the presence of α-ketoglutarate, a step involved in arginine metabolism. In Escherichia coli, the biosynthetic AcOAT also catalyzes the conversion of N-succinyl-l-2-amino-6-oxopimelate to N-succinyl-l,l-diaminopimelate, one of the steps in lysine biosynthesis. It is closely related to ornithine aminotransferase. AcOAT was cloned from Salmonella typhimurium and E. coli, overexpressed in E. coli and purified using Ni–NTA affinity column chromatography. The enzymes crystallized in the presence of gabaculine. Crystals of E. coli AcOAT (eAcOAT) only diffracted X-rays to 3.5 Å and were twinned. The crystals of S. typhimurium AcOAT (sAcOAT) diffracted to 1.9 Å and had a dimer in the asymmetric unit. The structure of sAcOAT was solved by the molecular-replacement method.

  18. A novel C-S lyase from the latex-producing plant Taraxacum brevicorniculatum displays alanine aminotransferase and l-cystine lyase activity.

    PubMed

    Munt, Oliver; Prüfer, Dirk; Schulze Gronover, Christian

    2013-01-01

    We isolated a novel pyridoxal-5-phosphate-dependent l-cystine lyase from the dandelion Taraxacum brevicorniculatum. Real time qPCR analysis showed that C-S lyase from Taraxacum brevicorniculatum (TbCSL) mRNA is expressed in all plant tissues, although at relatively low levels in the latex and pedicel. The 1251 bp TbCSL cDNA encodes a protein with a calculated molecular mass of 46,127 kDa. It is homologous to tyrosine and alanine aminotransferases (AlaATs) as well as to an Arabidopsis thaliana carbon-sulfur lyase (C-S lyase) (SUR1), which has a role in glucosinolate metabolism. TbCSL displayed in vitrol-cystine lyase and AlaAT activities of 4 and 19nkatmg(-1) protein, respectively. However, we detected no in vitro tyrosine aminotransferase (TyrAT) activity and RNAi knockdown of the enzyme had no effect on phenotype, showing that TbCSL substrates might be channeled into redundant pathways. TbCSL is in vivo localized in the cytosol and functions as a C-S lyase or an aminotransferase in planta, but the purified enzyme converts at least two substrates specifically, and can thus be utilized for further in vitro applications.

  19. Prognostic significance of preoperative aspartate aminotransferase to neutrophil ratio index in patients with hepatocellular carcinoma after hepatic resection

    PubMed Central

    Guo, Zhiyong; Pang, Hui; Chen, Dubo; Wang, Xiaoping; Ju, Weiqiang; Wang, Dongping; He, Xiaoshun; Hua, Yunpeng; Peng, Baogang

    2016-01-01

    Objectives Various inflammation-based prognostic scores have been associated with poor survival in patients with hepatocellular carcinoma (HCC), and neutrophils display important roles. However, few studies have illuminated the relationship between preoperative aspartate aminotransferase (AST) to neutrophil ratio index (ANRI) and poor prognosis of HCC. We aimed to clarify the prognostic value of ANRI and evaluate the ability of different inflammation-based prognostic scores such as ANRI, AST to lymphocyte ratio index (ALRI), AST to platelet count ratio index (APRI), neutrophil-lymphocyte ratio index (NLR), and platelet-lymphocyte ratio index (PLR). Methods Data were collected retrospectively from 303 patients who underwent curative resection for HCC. Preoperative ANRI, ALRI, APRI, NLR, PLR and clinico-pathological variables were analyzed. Univariate, multivariate and Kaplan-Meier analyses were performed to identify the predictive value of the above factors for disease-free survival (DFS) and overall survival (OS). Results ANRI was correlated with presence of HBsAg, AST, presence of cirrhosis, tumor size, PVTT, cancer of the liver Italian program (CLIP) score, recurrence. Univariate analysis showed ANRI, ALRI, APRI, NLR, PLR were significantly associated with DFS and OS in HCC patients with curative resection. After multivariate analysis, ANRI was demonstrated to be superior to ALRI, APRI, NLR, PLR, which were independently correlated with DFS and OS. Survival analysis showed that preoperative ANRI > 7.8 predicted poor prognosis of patients with HCC after hepatectomy. preoperative ANRI also showed different prognostic value in various subgroups of HCC. Furthermore, the predictive range was expanded by the combination of ANRI and NLR. Conclusions preoperative ANRI is an independent effective predictor of prognosis for patients with HCC, higher levels of ANRI predict poorer outcomes and the combining ANRI and NLR increases the prognostic accuracy of testing. PMID

  20. L,L-diaminopimelate aminotransferase (DapL): a putative target for the development of narrow-spectrum antibacterial compounds.

    PubMed

    Triassi, Alexander J; Wheatley, Matthew S; Savka, Michael A; Gan, Han Ming; Dobson, Renwick C J; Hudson, André O

    2014-01-01

    Despite the urgent need for sustained development of novel antibacterial compounds to combat the drastic rise in antibiotic resistant and emerging bacterial infections, only a few clinically relevant antibacterial drugs have been recently developed. One of the bottlenecks impeding the development of novel antibacterial compounds is the identification of new enzymatic targets. The nutritionally essential amino acid anabolic pathways, for example lysine biosynthesis, provide an opportunity to explore the development of antibacterial compounds, since human genomes do not possess the genes necessary to synthesize these amino acids de novo. The diaminopimelate (DAP)/lysine (lys) anabolic pathways are attractive targets for antibacterial development since the penultimate lys precursor meso-DAP (m-DAP) is a cross-linking amino acid in the peptidoglycan (PG) cell wall of most Gram-negative bacteria and lys plays a similar role in the PG of most Gram-positive bacteria, in addition to its role as one of the 20 proteogenic amino acids. The L,L-diaminopimelate aminotransferase (DapL) pathway was recently identified as a novel variant of the DAP/lys anabolic pathways. The DapL pathway has been identified in the pathogenic bacteria belonging to the genus; Chlamydia, Leptospira, and Treponema. The dapL gene has been identified in the genomes of 381 or approximately 13% of the 2771 bacteria that have been sequenced, annotated and reposited in the NCBI database, as of May 23, 2014. The narrow distribution of the DapL pathway in the bacterial domain provides an opportunity for the development and or discovery of narrow spectrum antibacterial compounds.

  1. Dose-Response Relationship between Alanine Aminotransferase Levels within the Reference Interval and Metabolic Syndrome in Chinese Adults

    PubMed Central

    Wu, Peipei; Chen, Qicai; Chen, Lili; Zhang, Pengpeng; Xiao, Juan; Chen, Xiaoxiao; Liu, Meng

    2017-01-01

    Purpose Elevation in serum alanine aminotransferase (ALT) levels is a biomarker for metabolic syndrome (MS); however, the relationship has not been fully investigated within the reference interval of ALT levels. Our objective was to explore the relationship between serum ALT levels within the reference interval and MS in Chinese adults. Materials and Methods This cross-sectional study included 16028 adults, who attended routine health check-ups at Shengli Oilfield Central Hospital from January 2006 to March 2012. The reference interval of serum ALT level was defined as less than 40 U/L. Logistic regression models and restricted cubic spline were used to evaluate the association of ALT with MS. Results The prevalence of MS in the total population was 13.7% (6.4% for females and 18.4% for males). Multiple logistic regression showed that ALT levels were positively associated with MS after adjustment for potential confounding factors. The odds ratio of MS in the top quartile was 4.830 [95% confidence interval (CI): 2.980–7.829] in females and 3.168 (95% CI: 2.649–3.790) in males, compared with the ALT levels in the bottom quartile. The restricted cubic spline models revealed a positive non-linear dose-response relationship between ALT levels and the risk of MS in women (p for nonlinearity was 0.0327), but a positive linear dose-response relationship in men (p for nonlinearity was 0.0659). Conclusion Serum ALT levels within the reference interval are positively associated with MS in a dose-response manner. Elevated ALT levels, even within the reference interval, may reflect early dysmetabolic changes. PMID:27873509

  2. Expression of mitochondrial branched-chain aminotransferase and α-keto-acid dehydrogenase in rat brain: implications for neurotransmitter metabolism

    PubMed Central

    Cole, Jeffrey T.; Sweatt, Andrew J.; Hutson, Susan M.

    2012-01-01

    In the brain, metabolism of the essential branched chain amino acids (BCAAs) leucine, isoleucine, and valine, is regulated in part by protein synthesis requirements. Excess BCAAs are catabolized or excreted. The first step in BCAA catabolism is catalyzed by the branched chain aminotransferase (BCAT) isozymes, mitochondrial BCATm and cytosolic BCATc. A product of this reaction, glutamate, is the major excitatory neurotransmitter and precursor of the major inhibitory neurotransmitter γ-aminobutyric acid (GABA). The BCATs are thought to participate in a α-keto-acid nitrogen shuttle that provides nitrogen for synthesis of glutamate from α-ketoglutarate. The branched-chain α-keto acid dehydrogenase enzyme complex (BCKDC) catalyzes the second, irreversible step in BCAA metabolism, which is oxidative decarboxylation of the branched-chain α-keto acid (BCKA) products of the BCAT reaction. Maple Syrup Urine Disease (MSUD) results from genetic defects in BCKDC, which leads to accumulation of toxic levels of BCAAs and BCKAs that result in brain swelling. Immunolocalization of BCATm and BCKDC in rats revealed that BCATm is present in astrocytes in white matter and in neuropil, while BCKDC is expressed only in neurons. BCATm appears uniformly distributed in astrocyte cell bodies throughout the brain. The segregation of BCATm to astrocytes and BCKDC to neurons provides further support for the existence of a BCAA-dependent glial-neuronal nitrogen shuttle since the data show that BCKAs produced by glial BCATm must be exported to neurons. Additionally, the neuronal localization of BCKDC suggests that MSUD is a neuronal defect involving insufficient oxidation of BCKAs, with secondary effects extending beyond the neuron. PMID:22654736

  3. TMEM199 Deficiency Is a Disorder of Golgi Homeostasis Characterized by Elevated Aminotransferases, Alkaline Phosphatase, and Cholesterol and Abnormal Glycosylation.

    PubMed

    Jansen, Jos C; Timal, Sharita; van Scherpenzeel, Monique; Michelakakis, Helen; Vicogne, Dorothée; Ashikov, Angel; Moraitou, Marina; Hoischen, Alexander; Huijben, Karin; Steenbergen, Gerry; van den Boogert, Marjolein A W; Porta, Francesco; Calvo, Pier Luigi; Mavrikou, Mersyni; Cenacchi, Giovanna; van den Bogaart, Geert; Salomon, Jody; Holleboom, Adriaan G; Rodenburg, Richard J; Drenth, Joost P H; Huynen, Martijn A; Wevers, Ron A; Morava, Eva; Foulquier, François; Veltman, Joris A; Lefeber, Dirk J

    2016-02-04

    Congenital disorders of glycosylation (CDGs) form a genetically and clinically heterogeneous group of diseases with aberrant protein glycosylation as a hallmark. A subgroup of CDGs can be attributed to disturbed Golgi homeostasis. However, identification of pathogenic variants is seriously complicated by the large number of proteins involved. As part of a strategy to identify human homologs of yeast proteins that are known to be involved in Golgi homeostasis, we identified uncharacterized transmembrane protein 199 (TMEM199, previously called C17orf32) as a human homolog of yeast V-ATPase assembly factor Vph2p (also known as Vma12p). Subsequently, we analyzed raw exome-sequencing data from families affected by genetically unsolved CDGs and identified four individuals with different mutations in TMEM199. The adolescent individuals presented with a mild phenotype of hepatic steatosis, elevated aminotransferases and alkaline phosphatase, and hypercholesterolemia, as well as low serum ceruloplasmin. Affected individuals showed abnormal N- and mucin-type O-glycosylation, and mass spectrometry indicated reduced incorporation of galactose and sialic acid, as seen in other Golgi homeostasis defects. Metabolic labeling of sialic acids in fibroblasts confirmed deficient Golgi glycosylation, which was restored by lentiviral transduction with wild-type TMEM199. V5-tagged TMEM199 localized with ERGIC and COPI markers in HeLa cells, and electron microscopy of a liver biopsy showed dilated organelles suggestive of the endoplasmic reticulum and Golgi apparatus. In conclusion, we have identified TMEM199 as a protein involved in Golgi homeostasis and show that TMEM199 deficiency results in a hepatic phenotype with abnormal glycosylation.

  4. Prevalence of elevated alanine-aminotransferase (ALT) among US adolescents and associated factors: NHANES 1999-2004

    PubMed Central

    Fraser, Abigail; Longnecker, Matthew P.; Lawlor, Debbie A

    2007-01-01

    Background & aims Non-alcoholic fatty liver disease (NAFLD) is a common cause of liver disease in children and adolescents. The majority of studies of NAFLD in children have been in select populations of the clinically obese. Study aims were to estimate the prevalence of elevated alanine-aminotransferase (ALT, as a marker of NAFLD) in a general contemporary adolescent population and to identify leading risk factors for ALT elevation (> 30 U/L). Methods We analysed data of adolescent participants (age 12-19, N=5586) in NHANES 1999-2004, a representative sample of the civilian non-institutionalized U.S population. Results The prevalence of elevated ALT (>30 U/L) was 7∙4% among white adolescents, 11∙5%, among Mexican Americans, and 6∙0%, among black adolescents. It was prevalent in 12∙4% of males compared to 3∙5% of females. Multivariable associations with elevated ALT were found for sex (OR male versus female = 7∙7, 95%CI: 3∙9, 15∙1), ethnicity (OR black versus white=0∙6, 95%CI: 0∙3, 1∙3; OR Mexican American versus white=1∙6, 95%CI: 1∙0, 2∙6), waist circumference (OR per 1 SD=1∙4, 95%CI: 1∙0, 2∙0), and fasting insulin (OR per 1 SD=1∙ 6, 95%CI: 1∙ 2, 2∙ 1). Age, C-reactive protein and triglycerides were also positively, and socio-economic position inversely associated with elevated ALT. The magnitude of associations with ALT was similar across ethnic groups. Conclusions ALT is associated with waist circumference and insulin resistance even in a young population. These characteristics could be utilized to identify adolescents who may benefit from screening for NAFLD, offering an opportunity to prevent disease progression at an early age. PMID:18054554

  5. Two mechanisms for putrescine-dependent transcriptional expression of the putrescine aminotransferase gene, ygjG, in Escherichia coli.

    PubMed

    Kim, Young-Sik; Shin, Hyun-Chul; Lee, Jong-Ho

    2014-09-01

    In this study, on evaluating the physiological function and mechanism of putrescine, we found that putrescine supplementation (1 mM) increases transcription of the putrescine aminotransferase gene, ygjG. Putrescine-dependent expression was confirmed by measuring β-galactosidase activity and with reverse transcription-polymerase chain reaction. To understand the role of putrescine in ygjG expression, we genetically characterized and found that a knockout mutation in an alternative sigma factor, rpoS, abolished putrescine-dependent ygjG-lacZ expression. In the rpoS mutant, RpoS overexpression complemented the mutant phenotype. However, RpoS overexpression induced ygjG-lacZ expression with putrescine supplementation but not without supplementation. We also found that the loss of putrescine-dependent ygjG-lacZ expression induced by rpoS was completely restored under nitrogen-starvation conditions. The putrescine-dependent expression of ygjG-lacZ under this condition was clearly dependent on another alternative sigma factor, rpoN, and its cognate activator ntrC. These results show that rpoS is required for putrescine-dependent ygjG-lacZ expression, but the effect of putrescine on this expression is not caused by simple modulation of RpoS synthesis. Putrescine-dependent expression of ygjG-lacZ was controlled by at least two sigma factors: rpoS under excess nitrogen conditions and rpoN under nitrogen-starvation conditions. These results suggest that putrescine plays an important role in the nitrogen regulation system.

  6. Diet and the frequency of the alanine:glyoxylate aminotransferase Pro11Leu polymorphism in different human populations.

    PubMed

    Caldwell, Elizabeth F; Mayor, Lianne R; Thomas, Mark G; Danpure, Christopher J

    2004-11-01

    The intermediary metabolic enzyme alanine:glyoxylate aminotransferase (AGT) contains a Pro11Leu polymorphism that decreases its catalytic activity by a factor of three and causes a small proportion to be mistargeted from its normal intracellular location in the peroxisomes to the mitochondria. These changes are predicted to have significant effects on the synthesis and excretion of the metabolic end-product oxalate and the deposition of insoluble calcium oxalate in the kidney and urinary tract. Based on the evolution of AGT targeting in mammals, we have previously hypothesised that this polymorphism would be advantageous for individuals who have a meat-rich diet, but disadvantageous for those who do not. If true, the frequency distribution of Pro11Leu in different extant human populations should have been shaped by their dietary history so that it should be more common in populations with predominantly meat-eating ancestral diets than it is in populations in which the ancestral diets were predominantly vegetarian. In the present study, we have determined frequency of Pro11Leu in 11 different human populations with divergent ancestral dietary lifestyles. We show that the Pro11Leu allelic frequency varies widely from 27.9% in the Saami, a population with a very meat-rich ancestral diet, to 2.3% in Chinese, who are likely to have had a more mixed ancestral diet. FST analysis shows that the differences in Pro11Leu frequency between some populations (particularly Saami vs Chinese) was very high when compared with neutral loci, suggesting that its frequency might have been shaped by dietary selection pressure.

  7. L,L-diaminopimelate aminotransferase (DapL): a putative target for the development of narrow-spectrum antibacterial compounds

    PubMed Central

    Triassi, Alexander J.; Wheatley, Matthew S.; Savka, Michael A.; Gan, Han Ming; Dobson, Renwick C. J.; Hudson, André O.

    2014-01-01

    Despite the urgent need for sustained development of novel antibacterial compounds to combat the drastic rise in antibiotic resistant and emerging bacterial infections, only a few clinically relevant antibacterial drugs have been recently developed. One of the bottlenecks impeding the development of novel antibacterial compounds is the identification of new enzymatic targets. The nutritionally essential amino acid anabolic pathways, for example lysine biosynthesis, provide an opportunity to explore the development of antibacterial compounds, since human genomes do not possess the genes necessary to synthesize these amino acids de novo. The diaminopimelate (DAP)/lysine (lys) anabolic pathways are attractive targets for antibacterial development since the penultimate lys precursor meso-DAP (m-DAP) is a cross-linking amino acid in the peptidoglycan (PG) cell wall of most Gram-negative bacteria and lys plays a similar role in the PG of most Gram-positive bacteria, in addition to its role as one of the 20 proteogenic amino acids. The L,L-diaminopimelate aminotransferase (DapL) pathway was recently identified as a novel variant of the DAP/lys anabolic pathways. The DapL pathway has been identified in the pathogenic bacteria belonging to the genus; Chlamydia, Leptospira, and Treponema. The dapL gene has been identified in the genomes of 381 or approximately 13% of the 2771 bacteria that have been sequenced, annotated and reposited in the NCBI database, as of May 23, 2014. The narrow distribution of the DapL pathway in the bacterial domain provides an opportunity for the development and or discovery of narrow spectrum antibacterial compounds. PMID:25309529

  8. Genetic variations in the alanine-glyoxylate aminotransferase 2 (AGXT2) gene and dimethylarginines levels in rheumatoid arthritis.

    PubMed

    Dimitroulas, Theodoros; Hodson, James; Panoulas, Vasileios F; Sandoo, Aamer; Smith, Jacqueline; Kitas, George

    2017-03-29

    Rheumatoid arthritis (RA) is associated with high rates of cardiovascular events mainly due to coronary and cerebrovascular atherosclerotic disease. Asymmetric (ADMA) and symmetric (SDMA) dimethylarginines are endogenous inhibitors of nitric oxide synthase and have been repeatedly linked with adverse cardiovascular outcomes in the general population and various disease settings. Alanine-glyoxylate aminotransferase 2 (AGTX2) is considered an alternative metabolic pathway contributing to the clearance of dimethylarginines in humans. The aim of the current study was to investigate the effect of specific AGXT-2 gene polymorphisms on circulating levels of ADMA or SDMA in patients with RA. Serum ADMA and SDMA levels were measured in 201 individuals with RA [median age: 67 years (IQR: 59-73), 155 females]. Two single nucleotide polymorphisms (SNPs) in the AGXT-2 gene-rs37369 and rs28305-were genotyped. Distributions of SDMA and ADMA were skewed, hence comparisons across the gene polymorphisms were performed using Kruskal-Wallis tests, and summarized using medians and interquartile ranges. Univariable analysis did not demonstrate a significant difference in the levels of SDMA or ADMA amongst the different genotypic groups of either rs37369AGXT2 (p = 0.800, 0.977) or rs28305AGXT2 (p = 0.463, 0.634). In multivariable analyses, ADMA levels were found to be significantly associated with erythrocyte sedimentation rate and estimated glomerular filtration rate, whilst SDMA levels were significantly associated with estimated glomerular filtration rate and quantitative insulin sensitivity check index. After adjustments for these factors, the relationship between the AGXT2 gene variants and both ADMA and SDMA remained non-significant. Our study in a well-characterized RA population did not show an association between serum concentrations of dimethylarginines and genetic variants of the AGXT2 gene.

  9. Identification of mutations associated with peroxisome-to-mitochondrion mistargeting of alanine/glyoxylate aminotransferase in primary hyperoxaluria type 1

    PubMed Central

    1990-01-01

    We have previously shown that in some patients with primary hyperoxaluria type 1 (PH1), disease is associated with mistargeting of the normally peroxisomal enzyme alanine/glyoxylate aminotransferase (AGT) to mitochondria (Danpure, C.J., P.J. Cooper, P.J. Wise, and P.R. Jennings. J. Cell Biol. 108:1345-1352). We have synthesized, amplified, cloned, and sequenced AGT cDNA from a PH1 patient with mitochondrial AGT (mAGT). This identified three point mutations that cause amino acid substitutions in the predicted AGT protein sequence. Using PCR and allele-specific oligonucleotide hybridization, a range of PH1 patients and controls were screened for these mutations. This revealed that all eight PH1 patients with mAGT carried at least one allele with the same three mutations. Two were homozygous for this allele and six were heterozygous. In at least three of the heterozygotes, it appeared that only the mutant allele was expressed. All three mutations were absent from PH1 patients lacking mAGT. One mutation encoding a Gly----Arg substitution at residue 170 was not found in any of the control individuals. However, the other two mutations, encoding Pro----Leu and Ile----Met substitutions at residues 11 and 340, respectively, cosegregated in the normal population at an allelic frequency of 5-10%. In an individual homozygous for this allele (substitutions at residues 11 and 340) only a small proportion of AGT appeared to be rerouted to mitochondria. It is suggested that the substitution at residue 11 generates an amphiphilic alpha-helix with characteristics similar to recognized mitochondrial targeting sequences, the full functional expression of which is dependent upon coexpression of the substitution at residue 170, which may induce defective peroxisomal import. PMID:1703535

  10. Characterization of the Branched-Chain Amino Acid Aminotransferase Enzyme Family in Tomato1[W][OA

    PubMed Central

    Maloney, Gregory S.; Kochevenko, Andrej; Tieman, Denise M.; Tohge, Takayuki; Krieger, Uri; Zamir, Dani; Taylor, Mark G.; Fernie, Alisdair R.; Klee, Harry J.

    2010-01-01

    Branched-chain amino acids (BCAAs) are synthesized in plants from branched-chain keto acids, but their metabolism is not completely understood. The interface of BCAA metabolism lies with branched-chain aminotransferases (BCAT) that catalyze both the last anabolic step and the first catabolic step. In this study, six BCAT genes from the cultivated tomato (Solanum lycopersicum) were identified and characterized. SlBCAT1, -2, -3, and -4 are expressed in multiple plant tissues, while SlBCAT5 and -6 were undetectable. SlBCAT1 and -2 are located in the mitochondria, SlBCAT3 and -4 are located in chloroplasts, while SlBCAT5 and -6 are located in the cytosol and vacuole, respectively. SlBCAT1, -2, -3, and -4 were able to restore growth of Escherichia coli BCAA auxotrophic cells, but SlBCAT1 and -2 were less effective than SlBCAT3 and -4 in growth restoration. All enzymes were active in the forward (BCAA synthesis) and reverse (branched-chain keto acid synthesis) reactions. SlBCAT3 and -4 exhibited a preference for the forward reaction, while SlBCAT1 and -2 were more active in the reverse reaction. While overexpression of SlBCAT1 or -3 in tomato fruit did not significantly alter amino acid levels, an expression quantitative trait locus on chromosome 3, associated with substantially higher expression of Solanum pennellii BCAT4, did significantly increase BCAA levels. Conversely, antisense-mediated reduction of SlBCAT1 resulted in higher levels of BCAAs. Together, these results support a model in which the mitochondrial SlBCAT1 and -2 function in BCAA catabolism while the chloroplastic SlBCAT3 and -4 function in BCAA synthesis. PMID:20435740

  11. ALT (Alanine Aminotransferase) Test

    MedlinePlus

    ... to help recognize heart or muscle injury. ALT values are often compared to the results of other tests such as alkaline phosphatase (ALP) , total protein , and bilirubin to help determine which form of liver disease is present. ALT is often used to monitor the treatment ...

  12. AST (Aspartate Aminotransferase) Test

    MedlinePlus

    PLEASE NOTE: Your web browser does not have JavaScript enabled. Unless you enable Javascript , your ability to navigate and access the features of this website will be limited. ... Proceeds from website advertising help sustain Lab Tests Online. AACC is a not-for-profit organization and does not endorse non-AACC products and services. Advertising & Sponsorship: ...

  13. Mechanism of substrate recognition and PLP-induced conformational changes in LL-diaminopimelate aminotransferase from Arabidopsis thaliana.

    PubMed

    Watanabe, Nobuhiko; Clay, Matthew D; van Belkum, Marco J; Cherney, Maia M; Vederas, John C; James, Michael N G

    2008-12-31

    LL-Diaminopimelate aminotransferase (LL-DAP-AT), a pyridoxal phosphate (PLP)-dependent enzyme in the lysine biosynthetic pathways of plants and Chlamydia, is a potential target for the development of herbicides or antibiotics. This homodimeric enzyme converts L-tetrahydrodipicolinic acid (THDP) directly to LL-DAP using L-glutamate as the source of the amino group. Earlier, we described the 3D structures of native and malate-bound LL-DAP-AT from Arabidopsis thaliana (AtDAP-AT). Seven additional crystal structures of AtDAP-AT and its variants are reported here as part of an investigation into the mechanism of substrate recognition and catalysis. Two structures are of AtDAP-AT with reduced external aldimine analogues: N-(5'-phosphopyridoxyl)-L-glutamate (PLP-Glu) and N-(5'-phosphopyridoxyl)- LL-Diaminopimelate (PLP-DAP) bound in the active site. Surprisingly, they reveal that both L-glutamate and LL-DAP are recognized in a very similar fashion by the same sets of amino acid residues; both molecules adopt twisted V-shaped conformations. With both substrates, the alpha-carboxylates are bound in a salt bridge with Arg404, whereas the distal carboxylates are recognized via hydrogen bonds to the well-conserved side chains of Tyr37, Tyr125 and Lys129. The distal C(epsilon) amino group of LL-DAP is specifically recognized by several non-covalent interactions with residues from the other subunit (Asn309*, Tyr94*, Gly95*, and Glu97* (Amino acid designators followed by an asterisk (*) indicate that the residues originate in the other subunit of the dimer)) and by three bound water molecules. Two catalytically inactive variants of AtDAP-AT were created via site-directed mutagenesis of the active site lysine (K270N and K270Q). The structures of these variants permitted the observation of the unreduced external aldimines of PLP with L-glutamate and with LL-DAP in the active site, and revealed differences in the torsion angle about the PLP-substrate bond. Lastly, an apo

  14. Mechanism of Substrate Recognition And PLP-Induced Conformational Changes in II-Diaminopimelate Aminotransferase From Arabidopsis Thaliana

    SciTech Connect

    Watanabe, N.; Clay, M.D.; Belkum, M.J.van; Cherney, M.M.; Vederas, J.C.; James, M.N.G.

    2009-05-26

    LL-Diaminopimelate aminotransferase (LL-DAP-AT), a pyridoxal phosphate (PLP)-dependent enzyme in the lysine biosynthetic pathways of plants and Chlamydia, is a potential target for the development of herbicides or antibiotics. This homodimeric enzyme converts L-tetrahydrodipicolinic acid (THDP) directly to LL-DAP using L-glutamate as the source of the amino group. Earlier, we described the 3D structures of native and malate-bound LL-DAP-AT from Arabidopsis thaliana (AtDAP-AT). Seven additional crystal structures of AtDAP-AT and its variants are reported here as part of an investigation into the mechanism of substrate recognition and catalysis. Two structures are of AtDAP-AT with reduced external aldimine analogues: N-(5'-phosphopyridoxyl)-L-glutamate (PLP-Glu) and N-(5'-phosphopyridoxyl)- LL-Diaminopimelate (PLP-DAP) bound in the active site. Surprisingly, they reveal that both L-glutamate and LL-DAP are recognized in a very similar fashion by the same sets of amino acid residues; both molecules adopt twisted V-shaped conformations. With both substrates, the {alpha}-carboxylates are bound in a salt bridge with Arg404, whereas the distal carboxylates are recognized via hydrogen bonds to the well-conserved side chains of Tyr37, Tyr125 and Lys129. The distal C{sup {var_epsilon}} amino group of LL-DAP is specifically recognized by several non-covalent interactions with residues from the other subunit (Asn309*, Tyr94*, Gly95*, and Glu97* (Amino acid designators followed by an asterisk (*) indicate that the residues originate in the other subunit of the dimer)) and by three bound water molecules. Two catalytically inactive variants of AtDAP-AT were created via site-directed mutagenesis of the active site lysine (K270N and K270Q). The structures of these variants permitted the observation of the unreduced external aldimines of PLP with L-glutamate and with LL-DAP in the active site, and revealed differences in the torsion angle about the PLP-substrate bond. Lastly, an apo

  15. Trihalomethane exposure and biomonitoring for the liver injury indicator, alanine aminotransferase, in the United States population (NHANES 1999–2006)

    PubMed Central

    Burch, James B.; Everson, Todd M.; Seth, Ratanesh K.; Wirth, Michael D.; Chatterjee, Saurabh

    2015-01-01

    Exposure to trihalomethanes (or THMs: chloroform, bromoform, bromodichloromethane, and dibromochloromethane [DBCM]) formed via drinking water disinfection has been associated with adverse reproductive outcomes and cancers of the digestive or genitourinary organs. However, few studies have examined potential associations between THMs and liver injury in humans, even though experimental studies suggest that these agents exert hepatotoxic effects, particularly among obese individuals. This study examined participants in the National Health and Nutrition Examination Survey (1999–2006, N = 2781) to test the hypothesis that THMs are associated with liver injury as assessed by alanine aminotransferase (ALT) activity in circulation. Effect modification by body mass index (BMI) or alcohol consumption also was examined. Associations between blood THM concentrations and ALT activity were assessed using unconditional multiple logistic regression to calculate prevalence odds ratios (ORs) with 95% confidence intervals (CIs) for exposure among cases with elevated ALT activity (men: >40 IU/L, women: >30 IU/L) relative to those with normal ALT, after adjustment for variables that may confound the relationship between ALT and THMs. Compared to controls, cases were 1.35 times more likely (95% CI: 1.02, 1.79) to have circulating DBCM concentrations exceeding median values in the population. There was little evidence for effect modification by BMI, although the association varied by alcohol consumption. Among non-drinkers, cases were more likely than controls to be exposed to DBCM (OR: 3.30, 95% CI: 1.37–7.90), bromoform (OR: 2.88, 95% CI: 1.21–6.81), or brominated THMs (OR: 4.00, 95% CI: 1.31–12.1), but no association was observed among participants with low, or moderate to heavy alcohol consumption. Total THM levels exceeding benchmark exposure limits continue to be reported both in the United States and globally. Results from this study suggest a need for further

  16. SAFETY study: Alanine aminotransferase cutoff values are set too high for reliable detection of pediatric chronic liver disease

    PubMed Central

    Schwimmer, Jeffrey B.; Dunn, Winston; Norman, Gregory J.; Pardee, Perrie E.; Middleton, Michael S.; Kerkar, Nanda; Sirlin, Claude B.

    2010-01-01

    Background & Aims The appropriate alanine aminotransferase (ALT) threshold value to use for diagnosis of chronic liver disease in children is unknown. We sought to develop sex-specific, biology-based, pediatric ALT thresholds. Methods The screening ALT for elevation in today’s youth (SAFETY) study collected observational data from acute care children’s hospitals, the national health and nutrition examination survey (NHANES, 1999–2006), overweight children with and without non-alcoholic fatty liver disease (NAFLD), and children with chronic Hepatitis B virus (HBV) or Hepatitis C virus (HCV) infections. The study compared the sensitivity and specificity of ALT thresholds currently used by children’s hospitals versus study-derived, sex-specific, biology-based, ALT thresholds for detecting children with NAFLD, HCV, or HBV. Results The median upper limit of ALT at children’s hospitals was 53 U/L (range, 30–90). The 95th percentile levels for ALT in healthy weight, metabolically normal, liver disease-free, NHANES pediatric participants were 25.8 U/L (boys) and 22.1 U/L (girls). The concordance statistics of these NHANES-derived thresholds for liver disease detection were 0.85 (95% confidence interval [CI] 0.74–0.96) in boys and 0.91 (95% CI 0.83–0.99) in girls for NAFLD, 0.80 (95% CI 0.70–0.91) in boys and 0.79 (95% CI 0.69–0.89) in girls for HBV, and 0.86 (95% CI 0.77–0.95) in boys and 0.84 (95% CI 0.75–0.93) in girls for HCV. Using current children’s hospitals ALT thresholds, the median sensitivity for detection of NAFLD, HBV, and HCV ranged from 32% to 48%; median specificity was 92% (boys) and 96% (girls). Using NHANES-derived thresholds, the sensitivities were 72% (boys) and 82% (girls); specificities were 79% (boys) and 85% (girls). Conclusions The upper limit of ALT used in children’s hospitals varies widely and is set too high to reliably detect chronic liver disease. Biology-based thresholds provide higher sensitivity and only

  17. Structural and Functional Characterization of PseC, an Aminotransferase Involved in the Biosynthesis of Pseudaminic Acid, an Essential Flagellar Modification in Helicobacter Pylori

    SciTech Connect

    Schoenhofen,I.; Lunin, V.; Julien, J.; Li, Y.; Ajamian, E.; Matte, A.; Cygler, M.; Brisson, J.; Aubry, A.; et al.

    2006-01-01

    Helicobacter pylori flagellin is heavily glycosylated with the novel sialic acid-like nonulosonate, pseudaminic acid (Pse). The glycosylation process is essential for assembly of functional flagellar filaments and consequent bacterial motility. As motility is a key virulence factor for this and other important pathogens, the Pse biosynthetic pathway offers potential for novel therapeutic targets. From recent NMR analyses, we determined that the conversion of UDP-a-D-GlcNAc to the central intermediate in the pathway, UDP-4-amino-4,6-dideoxy-{beta}-L-AltNAc, proceeds by formation of UDP-2-acetamido-2,6-dideoxy-{beta}-L-arabino-4-hexulose by the dehydratase/epimerase PseB (HP0840) followed with amino transfer by the aminotransferase, PseC (HP0366). The central role of PseC in the H. pylori Pse biosynthetic pathway prompted us to determine crystal structures of the native protein, its complexes with pyridoxal phosphate alone and in combination with the UDP-4-amino-4,6-dideoxy-{beta}-L-AltNAc product, the latter being converted to the external aldimine form in the enzyme's active site. In the binding site, the AltNAc sugar ring adopts a 4C1 chair conformation which is different from the predominant 1C4 form found in solution. The enzyme forms a homodimer where each monomer contributes to the active site, and these structures have permitted the identification of key residues involved in stabilization, and possibly catalysis, of the {beta}-L-arabino intermediate during the amino transfer reaction. The essential role of Lys183 in the catalytic event was confirmed by site-directed mutagenesis. This work presents for the first time a nucleotide-sugar aminotransferase co-crystallized with its natural ligand, and in conjunction with the recent functional characterization of this enzyme, will assist in elucidating the aminotransferase reaction mechanism within the Pse biosynthetic pathway.

  18. Structural Analysis of QdtB, an Aminotransferase Required for the Biosynthesis of dTDP-3-acetamido-3,6-dideoxy-[alpha]-D-glucose

    SciTech Connect

    Thoden, James B.; Schaffer, Christina; Messner, Paul; Holden, Hazel M.

    2009-05-21

    3-Acetamido-3,6-dideoxy-{alpha}-D-glucose or Quip3NAc is an unusual deoxyamino sugar found in the O-antigens of some Gram-negative bacteria and in the S-layers of Gram-positive bacteria. It is synthesized in these organisms as a dTDP-linked sugar via the action of five enzymes. The focus of this investigation is on QdtB from Thermoanaerobacterium thermosaccharolyticum E207-71, a PLP-dependent aminotransferase that catalyzes the penultimate step in the production of dTDP-Quip3NAc. For this analysis, the enzyme was crystallized in the presence of its product, dTDP-Quip3N, and the structure was solved and refined to 2.15 {angstrom} resolution. QdtB is a dimer, and its overall fold places it into the well-characterized aspartate aminotransferase superfamily. Electron density corresponding to the bound product reveals the presence of a Schiff base between C-4' of the PLP cofactor and the amino nitrogen of the sugar. Those amino acid side chains involved in binding the dTDP-sugar into the active site include Tyr 183, His 309, and Tyr 310 from subunit 1 and Lys 219 from subunit 2. Notably there is a decided lack of interactions between the pyranosyl C-4' hydroxyl of the dTDP-sugar and the protein. In keeping with this observation, we show that QdtB can also turn over dTDP-3-acetamido-3,6-dideoxy-{alpha}-D-galactose. This investigation represents the first structural analysis of a sugar-modifying aminotransferase with a bound product in its active site that functions at the C-3' rather than the C-4' position of the hexose.

  19. Differential response of orthologous L,L-diaminopimelate aminotransferases (DapL) to enzyme inhibitory antibiotic lead compounds.

    PubMed

    McKinnie, Shaun M K; Rodriguez-Lopez, Eva M; Vederas, John C; Crowther, Jennifer M; Suzuki, Hironori; Dobson, Renwick C J; Leustek, Thomas; Triassi, Alexander J; Wheatley, Matthew S; Hudson, André O

    2014-01-01

    L,L-Diaminopimelate aminotransferase (DapL) is an enzyme required for the biosynthesis of meso-diaminopimelate (m-DAP) and L-lysine (Lys) in some bacteria and photosynthetic organisms. m-DAP and Lys are both involved in the synthesis of peptidoglycan (PG) and protein synthesis. DapL is found in specific eubacterial and archaeal lineages, in particular in several groups of pathogenic bacteria such as Leptospira interrogans (LiDapL), the soil/water bacterium Verrucomicrobium spinosum (VsDapL) and the alga Chlamydomonas reinhardtii (CrDapL). Here we present the first comprehensive inhibition study comparing the kinetic activity of DapL orthologs using previously active small molecule inhibitors formerly identified in a screen with the DapL of Arabidopsis thaliana (AtDapL), a flowering plant. Each inhibitor is derived from one of four classes with different central structural moieties: a hydrazide, a rhodanine, a barbiturate, or a thiobarbituate functionality. The results show that all five compounds tested were effective at inhibiting the DapL orthologs. LiDapL and AtDapL showed similar patterns of inhibition across the inhibitor series, whereas the VsDapL and CrDapL inhibition patterns were different from that of LiDapL and AtDapL. CrDapL was found to be insensitive to the hydrazide (IC₅₀ >200 μM). VsDapL was found to be the most sensitive to the barbiturate and thiobarbiturate containing inhibitors (IC₅₀ ∼5 μM). Taken together, the data shows that the homologs have differing sensitivities to the inhibitors with IC₅₀ values ranging from 4.7 to 250 μM. In an attempt to understand the basis for these differences the four enzymes were modeled based on the known structure of AtDapL. Overall, it was found that the enzyme active sites were conserved, although the second shell of residues close to the active site were not. We conclude from this that the altered binding patterns seen in the inhibition studies may be a consequence of the inhibitors forming

  20. Meta-analysis of the influence of TM6SF2 E167K variant on Plasma Concentration of Aminotransferases across different Populations and Diverse Liver Phenotypes

    PubMed Central

    Sookoian, Silvia; Pirola, Carlos J.

    2016-01-01

    A nonsynonymous E167K (rs58542926 C/T) variant in TM6SF2 gene was recently associated with nonalcoholic fatty liver disease (NAFLD). We explored the association between E167K and plasma concentrations of alanine (ALT) and aspartate (AST) aminotransferases through a meta-analysis. We also estimated the strength of the effect across diverse liver phenotypes, including NAFLD and chronic viral hepatitis; fourteen studies were included. We found that ALT (p = 3.2 × 10−6, n = 94,414) and AST (p = 0007, n = 93,809) levels were significantly associated with rs58542926 in NAFLD. By contrast, rs58542926 was not associated with either ALT (p = 0.24, n = 4187) or AST (p = 0.17, n = 2678) levels in four studies on chronic hepatitis. In conclusion, the results of the pooled estimates in patients with NAFLD showed that carriers of the T allele (EK + KK), when compared with homozygous subjects for the C allele (EE genotype) have increased levels of aminotransferases; however, this increase represents –2.5 (9.8%) and 1.2 (5%) IU/L of ALT and AST respectively, which is fairly small compared with the large effect of PNPLA3- rs738409-G allele that is associated with a –28% increase in serum ALT. PMID:27278285

  1. Corticosterone, cortisol, triglycerides, aspartate aminotransferase and uric acid plasma concentrations during foie gras production in male mule ducks (Anas platyrhynchos × Cairina moschata).

    PubMed

    Flament, A; Delleur, V; Poulipoulis, A; Marlier, D

    2012-01-01

    1. Corticosterone, cortisol, triglycerides, aspartate aminotransferase (AST) and uric acid (UA) plasma concentration were measured at 8 (7 days after group housing), 12 (after 7 days of force feeding) and 13 weeks of age (at slaughter after 12 days of force feeding), and 45 min after an adrenocorticotrophic hormone (ACTH) stimulation test at 8 weeks of age in 12 male mule ducks in an on-farm experiment. 2. No significant increase of corticosterone was found during the force-feeding period compared with the concentration after housing. 3. Comparison of corticosterone and cortisol values indicates that cortisol can be considered as a reliable acute stress indicator in future routine examinations. 4. Plasma concentrations of triglycerides and aspartate aminotransferase increased progressively from pre-force feeding period to slaughtering. 5. Plasma concentrations of uric acid increased from the start at 8 weeks of age to the mid-force feeding period but no difference was noticed between the mid-force feeding period and slaughtering. 6. It is concluded that acute stress induced by force-feeding is similar at the beginning and end of the commercial production of foie gras.

  2. BarR, an Lrp-type transcription factor in Sulfolobus acidocaldarius, regulates an aminotransferase gene in a β-alanine responsive manner.

    PubMed

    Liu, Han; Orell, Alvaro; Maes, Dominique; van Wolferen, Marleen; Lindås, Ann-Christin; Bernander, Rolf; Albers, Sonja-Verena; Charlier, Daniel; Peeters, Eveline

    2014-05-01

    In archaea, nothing is known about the β-alanine degradation pathway or its regulation. In this work, we identify and characterize BarR, a novel Lrp-like transcription factor and the first one that has a non-proteinogenic amino acid ligand. BarR is conserved in Sulfolobus acidocaldarius and Sulfolobus tokodaii and is located in a divergent operon with a gene predicted to encode β-alanine aminotransferase. Deletion of barR resulted in a reduced exponential growth rate in the presence of β-alanine. Furthermore, qRT-PCR and promoter activity assays demonstrated that BarR activates the expression of the adjacent aminotransferase gene, but only upon β-alanine supplementation. In contrast, auto-activation proved to be β-alanine independent. Heterologously produced BarR is an octamer in solution and forms a single complex by interacting with multiple sites in the 170 bp long intergenic region separating the divergently transcribed genes. In vitro, DNA binding is specifically responsive to β-alanine and site-mutant analyses indicated that β-alanine directly interacts with the ligand-binding pocket. Altogether, this work contributes to the growing body of evidence that in archaea, Lrp-like transcription factors have physiological roles that go beyond the regulation of α-amino acid metabolism.

  3. Characterization of tryptophan aminotransferase 1 of Malassezia furfur, the key enzyme in the production of indolic compounds by M. furfur.

    PubMed

    Preuss, Janina; Hort, Wiebke; Lang, Sarah; Netsch, Anette; Rahlfs, Stefan; Lochnit, Günter; Jortzik, Esther; Becker, Katja; Mayser, Peter A

    2013-11-01

    Malassezia yeasts are responsible for the widely distributed skin disease Pityriasis versicolor (PV), which is characterized by a hyper- or hypopigmentation of affected skin areas. For Malassezia furfur, it has been shown that pigment production relies on tryptophan metabolism. A tryptophan aminotransferase was found to catalyse the initial catalytic step in pigment formation in the model organism Ustilago maydis. Here, we describe the sequence determination, recombinant production and biochemical characterization of tryptophan aminotransferase MfTam1 from M. furfur. The enzyme catalyses the transamination from l-tryptophan to keto acids such as α-ketoglutarate with Km values for both substrates in the low millimolar range. Furthermore, MfTam1 presents a temperature optimum at 40°C and a pH optimum at 8.0. MfTam1 activity is highly dependent on pyridoxal phosphate (PLP), whereas compounds interfering with PLP, such as cycloserine (CS) and aminooxyacetate, inhibit the MfTam1 reaction. CS is known to reverse hyperpigmentation in PV. Thus, the results of the present study give a deeper insight into the role of MfTam1 in PV pathogenesis and as potential target for the development of novel PV therapeutics.

  4. Crystal structure of LL-diaminopimelate aminotransferase from Arabidopsis thaliana: a recently discovered enzyme in the biosynthesis of L-lysine by plants and Chlamydia.

    PubMed

    Watanabe, Nobuhiko; Cherney, Maia M; van Belkum, Marco J; Marcus, Sandra L; Flegel, Mitchel D; Clay, Matthew D; Deyholos, Michael K; Vederas, John C; James, Michael N G

    2007-08-17

    The essential biosynthetic pathway to l-Lysine in bacteria and plants is an attractive target for the development of new antibiotics or herbicides because it is absent in humans, who must acquire this amino acid in their diet. Plants use a shortcut of a bacterial pathway to l-Lysine in which the pyridoxal-5'-phosphate (PLP)-dependent enzyme ll-diaminopimelate aminotransferase (LL-DAP-AT) transforms l-tetrahydrodipicolinic acid (L-THDP) directly to LL-DAP. In addition, LL-DAP-AT was recently found in Chlamydia sp., suggesting that inhibitors of this enzyme may also be effective against such organisms. In order to understand the mechanism of this enzyme and to assist in the design of inhibitors, the three-dimensional crystal structure of LL-DAP-AT was determined at 1.95 A resolution. The cDNA sequence of LL-DAP-AT from Arabidopsis thaliana (AtDAP-AT) was optimized for expression in bacteria and cloned in Escherichia coli without its leader sequence but with a C-terminal hexahistidine affinity tag to aid protein purification. The structure of AtDAP-AT was determined using the multiple-wavelength anomalous dispersion (MAD) method with a seleno-methionine derivative. AtDAP-AT is active as a homodimer with each subunit having PLP in the active site. It belongs to the family of type I fold PLP-dependent enzymes. Comparison of the active site residues of AtDAP-AT and aspartate aminotransferases revealed that the PLP binding residues in AtDAP-AT are well conserved in both enzymes. However, Glu97* and Asn309* in the active site of AtDAP-AT are not found at similar positions in aspartate aminotransferases, suggesting that specific substrate recognition may require these residues from the other monomer. A malate-bound structure of AtDAP-AT allowed LL-DAP and L-glutamate to be modelled into the active site. These initial three-dimensional structures of LL-DAP-AT provide insight into its substrate specificity and catalytic mechanism.

  5. Crystal Structure of Ll-Diaminopimelate Aminotransferase From 'Arabidopsis Thaliana': a Recently-Discovered Enzyme in the Biosynthesis of L-Lysine By Plants And 'Chlamydia'

    SciTech Connect

    Watanabe, N.; Cherney, M.M.; van Belkum, M.J.; Marcus, S.L.; Flegel, M.D.; Clay, M.D.; Deyholos, M.K.; Vederas, J.C.; James, M.N.G.

    2007-07-13

    The essential biosynthetic pathway to l-Lysine in bacteria and plants is an attractive target for the development of new antibiotics or herbicides because it is absent in humans, who must acquire this amino acid in their diet. Plants use a shortcut of a bacterial pathway to l-Lysine in which the pyridoxal-5-phosphate (PLP)-dependent enzyme ll-diaminopimelate aminotransferase (LL-DAP-AT) transforms l-tetrahydrodipicolinic acid (L-THDP) directly to LL-DAP. In addition, LL-DAP-AT was recently found in Chlamydia sp., suggesting that inhibitors of this enzyme may also be effective against such organisms. In order to understand the mechanism of this enzyme and to assist in the design of inhibitors, the three-dimensional crystal structure of LL-DAP-AT was determined at 1.95 Angstroms resolution. The cDNA sequence of LL-DAP-AT from Arabidopsis thaliana (AtDAP-AT) was optimized for expression in bacteria and cloned in Escherichia coli without its leader sequence but with a C-terminal hexahistidine affinity tag to aid protein purification. The structure of AtDAP-AT was determined using the multiple-wavelength anomalous dispersion (MAD) method with a seleno-methionine derivative. AtDAP-AT is active as a homodimer with each subunit having PLP in the active site. It belongs to the family of type I fold PLP-dependent enzymes. Comparison of the active site residues of AtDAP-AT and aspartate aminotransferases revealed that the PLP binding residues in AtDAP-AT are well conserved in both enzymes. However, Glu97* and Asn309* in the active site of AtDAP-AT are not found at similar positions in aspartate aminotransferases, suggesting that specific substrate recognition may require these residues from the other monomer. A malate-bound structure of AtDAP-AT allowed LL-DAP and L-glutamate to be modeled into the active site. These initial three-dimensional structures of LL-DAP-AT provide insight into its substrate specificity and catalytic mechanism.

  6. Isolation and characterization of a gene from Trypanosoma cruzi encoding a 46-kilodalton protein with homology to human and rat tyrosine aminotransferase.

    PubMed

    Bontempi, E J; Búa, J; Aslund, L; Porcel, B; Segura, E L; Henriksson, J; Orn, A; Pettersson, U; Ruiz, A M

    1993-06-01

    The complete sequence of a gene encoding a 46-kDa protein of Trypanosoma cruzi is presented. The first ATG complies with the consensus sequence for initiation of translation. A single band of 2 kb was highlighted by hybridizing a probe from the 46-kDa protein gene to a Northern filter containing total T. cruzi RNA. The gene is present in 50-80 copies per cell and most of them are contained in 2 tandem arrays on large T. cruzi chromosomes (> 2000 kb). A strong homology with rat and human tyrosine aminotransferase was detected. Homology with a Trypanosoma brucei retrotransposon was found in the nonsense strand of the intergenic region.

  7. Docking and quantitative structure-activity relationship studies for sulfonyl hydrazides as inhibitors of cytosolic human branched-chain amino acid aminotransferase.

    PubMed

    Caballero, Julio; Vergara-Jaque, Ariela; Fernández, Michael; Coll, Deysma

    2009-11-01

    We have performed the docking of sulfonyl hydrazides complexed with cytosolic branched-chain amino acid aminotransferase (BCATc) to study the orientations and preferred active conformations of these inhibitors. The study was conducted on a selected set of 20 compounds with variation in structure and activity. In addition, the predicted inhibitor concentration (IC(50)) of the sulfonyl hydrazides as BCAT inhibitors were obtained by a quantitative structure-activity relationship (QSAR) method using three-dimensional (3D) vectors. We found that three-dimensional molecule representation of structures based on electron diffraction (3D-MoRSE) scheme contains the most relevant information related to the studied activity. The statistical parameters [cross-validate correlation coefficient (Q(2) = 0.796) and fitted correlation coefficient (R(2) = 0.899)] validated the quality of the 3D-MoRSE predictive model for 16 compounds. Additionally, this model adequately predicted four compounds that were not included in the training set.

  8. Functional roles of a predicted branched chain aminotransferase encoded by the LkBAT1 gene of the yeast Lachancea kluyveri.

    PubMed

    Montalvo-Arredondo, Javier; Jiménez-Benítez, Ángel; Colón-González, Maritrini; González-Flores, James; Flores-Villegas, Mirelle; González, Alicia; Riego-Ruiz, Lina

    2015-12-01

    Branched chain amino acid aminotransferases (BCATs) catalyze the last step of the biosynthesis and the first step of the catabolism of branched chain amino acids. In Saccharomyces cerevisiae, BCATs are encoded by the ScBAT1 and ScBAT2 paralogous genes. Analysis of Lachancea kluyveri genome sequence, allowed the identification of the LkBAT1 locus, which could presumably encode a BCAT. A second unlinked locus (LkBAT1bis), exhibiting sequence similarity to LkBAT1 was also identified. To determine the function of these putative BCATs, L. kluyveri mutant strains lacking LkBAT1, LkBAT1bis or both genes were generated and tested for VIL metabolism. LkBat1 displayed branched chain aminotransferase activity and is required for VIL biosynthesis and catabolism. However, Lkbat1Δ mutant is a valine and isoleucine auxotroph and a leucine bradytroph indicating that L. kluyveri harbors an alternative enzyme(s) involved in leucine biosynthesis. Additionally, heterologous reciprocal gene complementation between S. cerevisiae and L. kluyveri orthologous LkBAT1, ScBAT1 and ScBAT2 genes, confirmed that the mitochondrial LkBat1 functions as BCAT in S. cerevisiae, restoring wild type phenotype to the ScBAT1 null mutant. Conversely, LkBAT1bis did not display a role in BCAAs metabolism. However, when ethanol was used as carbon source, deletion of LkBAT1bis in an Lkbat1Δ null strain resulted in an extended 'lag' growth phase, pointing to a potential function of LkBAT1 and LkBAT1bis in the aerobic metabolism of L. kluyveri. These results confirm the BCAT function of LkBAT1 in L. kluyveri, and further support the proposition that the BCAT function in ancestral-type yeasts has been distributed in the two paralogous genes present in S. cerevisiae.

  9. l,l-diaminopimelate aminotransferase, a trans-kingdom enzyme shared by Chlamydia and plants for synthesis of diaminopimelate/lysine

    PubMed Central

    McCoy, Andrea J.; Adams, Nancy E.; Hudson, André O.; Gilvarg, Charles; Leustek, Thomas; Maurelli, Anthony T.

    2006-01-01

    The synthesis of meso-diaminopimelic acid (m-DAP) in bacteria is essential for both peptidoglycan and lysine biosynthesis. From genome sequencing data, it was unclear how bacteria of the Chlamydiales order would synthesize m-DAP in the absence of dapD, dapC, and dapE, which are missing from the genome. Here, we assessed the biochemical capacity of Chlamydia trachomatis serovar L2 to synthesize m-DAP. Expression of the chlamydial asd, dapB, and dapF genes in the respective Escherichia coli m-DAP auxotrophic mutants restored the mutants to DAP prototrophy. Screening of a C. trachomatis genomic library in an E. coli ΔdapD DAP auxotroph identified ct390 as encoding an enzyme that restored growth to the Escherichia coli mutant. ct390 also was able to complement an E. coli ΔdapD ΔdapE, but not a ΔdapD ΔdapF mutant, providing genetic evidence that it encodes an aminotransferase that may directly convert tetrahydrodipicolinate to l,l-diaminopimelic acid. This hypothesis was supported by in vitro kinetic analysis of the CT390 protein and the fact that similar properties were demonstrated for the Protochlamydia amoebophila homologue, PC0685. In vivo, the C. trachomatis m-DAP synthesis genes are expressed as early as 8 h after infection. An aminotransferase activity analogous to CT390 recently has been characterized in plants and cyanobacteria. This previously undescribed pathway for m-DAP synthesis supports an evolutionary relationship among the chlamydiae, cyanobacteria, and plants and strengthens the argument that chlamydiae synthesize a cell wall despite the inability of efforts to date to detect peptidoglycan in these organisms. PMID:17093042

  10. CaAlaAT1 catalyzes the alanine: 2-oxoglutarate aminotransferase reaction during the resistance response against Tobacco mosaic virus in hot pepper.

    PubMed

    Kim, Ki-Jeong; Park, Chang-Jin; An, Jong-Min; Ham, Byung-Kook; Lee, Boo-Ja; Paek, Kyung-Hee

    2005-08-01

    Hot pepper (Capsicum annuum L. cv. Bugang) plants exhibit a hypersensitive response (HR) upon infection by Tobacco mosaic virus (TMV) pathotype P0. To elucidate molecular mechanism that underlies this resistance, hot pepper cv. Bugang leaves were inoculated with TMV-P0 and genes specifically up-regulated during the HR were isolated by differential screening. One of the clones, CaAlaAT1 encoding a putative alanine aminotransferase (EC 2.6.1.2) exhibited organ-specific expression pattern and the transcript accumulated abundantly in red (ripe) fruit tissues. CaAlaAT1 transcript was also induced in older leaves during senescence. The expression of CaAlaAT1 gene was increased in the incompatible interaction with TMV-P0 but was not in the compatible interaction with TMV-P1.2. When a strain of Xanthomonas campestris pv. vesicatoria (Xcv) carrying an AvrBs2 gene was infiltrated into the leaves of a pepper cv. ECW 20R carrying Bs2 resistance gene, a marked induction and maintenance of CaAlaAT1 gene expression was observed. The expression of CaAlaAT1 gene was triggered by salicylic acid (SA) and ethylene but not by methyl jasmonate (MeJA). CaAlaAT1 seemed to be localized mostly at the cytosol from the polyethylene glycol (PEG)-mediated transformation experiment. CaAlaAT1 seemed to catalyze alanine: 2-oxoglutarate aminotransferase (AKT) reaction, which was a main activity among the four activities in vitro, during the resistance response against TMV in hot pepper. These results suggest that CaAlaAT1, a protein known to be involved in metabolic reactions, might be one of the components in the plant's defense signal pathway against pathogens.

  11. The enzymology of alanine aminotransferase (AlaAT) isoforms from Hordeum vulgare and other organisms, and the HvAlaAT crystal structure.

    PubMed

    Duff, Stephen M G; Rydel, Timothy J; McClerren, Amanda L; Zhang, Wenlan; Li, Jimmy Y; Sturman, Eric J; Halls, Coralie; Chen, Songyang; Zeng, Jiamin; Peng, Jiexin; Kretzler, Crystal N; Evdokimov, Artem

    2012-12-01

    In this paper we describe the expression, purification, kinetics and biophysical characterization of alanine aminotransferase (AlaAT) from the barley plant (Hordeum vulgare). This dimeric PLP-dependent enzyme is a pivotal element of several key metabolic pathways from nitrogen assimilation to carbon metabolism, and its introduction into transgenic plants results in increased yield. The enzyme exhibits a bi-bi ping-pong reaction mechanism with a K(m) for alanine, 2-oxoglutarate, glutamate and pyruvate of 3.8, 0.3, 0.8 and 0.2 mM, respectively. Barley AlaAT catalyzes the forward (alanine-forming) reaction with a k(cat) of 25.6 s(-1), the reverse (glutamate-forming) reaction with k(cat) of 12.1 s(-1) and an equilibrium constant of ~0.5. The enzyme is also able to utilize aspartate and oxaloacetate with ~10% efficiency as compared to the native substrates, which makes it much more specific than related bacterial/archaeal enzymes (that also have lower K(m) values). We have crystallized barley AlaAT in complex with PLP and l-cycloserine and solved the structure of this complex at 2.7 Å resolution. This is the first example of a plant AlaAT structure, and it reveals a canonical aminotransferase fold similar to structures of the Thermotoga maritima, Pyrococcus furiosus, and human enzymes. This structure bridges our structural understanding of AlaAT mechanism between three kingdoms of life and allows us to shed some light on the specifics of the catalysis performed by these proteins.

  12. [The effect of diet ratio of polyunsaturated fatty acids of omega-3 and omega-6 families on activity of aminotransferases and gamma-glutamyltransferase in rat blood serum].

    PubMed

    Ketsa, O V; Marchenko, M M

    2014-01-01

    The effect of diet fat compositions with various ratio of omega-3 and omega-6 polyunsaturated fatty acids (PUFAs) on alanine aminotransferase (ALT), aspartate aminotransferase (AST) and gamma-glutamyltransferase (GGT) activities in blood serum of 45 white mongrel rats weighing 90-110 g (9 animals in group) has been investigated. Fat components in the semi-synthetic diet, compiled on the basis of AIN-93 diet, and sources of omega-6 and omega-3 PUFA were presented by sunflower oil, soybean oil and fish oil. It has been shown that four-week inclusion of linoleic acid (LA) and alpha-linolenic acid (alpha-LNA) in a ratio of 7:1 into the diet (soybean oil) as well as use of only omega-6 PUFA (sunflower oil) has lead to an increase in the activity of ALT and GGT in rat blood serum compared to control animals treated with the complex of linolenic, eicosapentaenoic (EPA) and docosahexaenoic (DHA) acid through the mixture of sunflower oil and fish oil (9:1) with the ratio of omega-6 and omega-3 PUFA 7:1. Along with this, the AST:ALT ratio (de Ritis ratio) was lower (p < 0.05) as compared with the control group of rat, amounting respectively 0.92 +/- 0.08 and 0.79 +/- 0.12 vs 1.26 +/- 0.10. The use of high doses of omega-3 fatty acids (600 mg EPA and 400 mg DHA per kg of animal weight per day coming through fish oil) did not affect the activity of ALT and GGT, but increased AST serum activity (0.47 +/- 0.04 micromoles/min per mg protein) and the de Ritis ratio (2.53 +/- 0.23). The diet deprived with fat increased enzyme activity of ALT, AST and GGT in rat blood serum.

  13. Relationship Between Hepatic Steatosis and the Elevation of Aminotransferases in HBV-Infected Patients With HBe-Antigen Negativity and a Low Viral Load

    PubMed Central

    Enomoto, Hirayuki; Aizawa, Nobuhiro; Nishikawa, Hiroki; Ikeda, Naoto; Sakai, Yoshiyuki; Takata, Ryo; Hasegawa, Kunihiro; Nakano, Chikage; Nishimura, Takashi; Yoh, Kazunori; Ishii, Akio; Takashima, Tomoyuki; Iwata, Yoshinori; Iijima, Hiroko; Nishiguchi, Shuhei

    2016-01-01

    Abstract Nonalcoholic fatty liver disease has been suggested to be associated with alanine aminotransferase (ALT) elevation in hepatitis B virus (HBV)-infected patients with HBe antigen (HBeAg)-negativity and a low HBV-DNA level. However, few studies have evaluated the association according to histological findings of the liver. Among a total of 198 HBV-infected patients who received a percutaneous liver biopsy, we studied the histological and laboratory findings of HBeAg-negative patients without receiving nucleoside/nucleotide analogues treatment (N = 70) in order to evaluate whether hepatic steatosis and its related metabolic disorders were associated with an elevation in ALT levels in HBeAg-negative patients. In HBeAg-negative patients with a high serum HBV-DNA level (≥2000 IU/mL), the level of HBV-DNA was the only significant factor related to ALT elevation. However, in HBeAg-negative patients with a low HBV-DNA level, the serum ferritin level, and histologically observed hepatic steatosis were significantly associated factors with ALT elevation. When we evaluated 2 metabolic variables (serum ferritin and fasting insulin) that are suggested to be relevant to the presence of progressive disease in Japanese patients, we found that the rate of metabolic disorders was significantly higher among patients with a high ALT level and a low HBV-DNA level than it was among those with other conditions. The triglyceride level and the frequency of moderate or severe hepatic steatosis were significantly higher in patients with a low HBV-DNA level than in those with a high HBV-DNA level. Histologically proven hepatic steatosis and its related metabolic disorders are suggested to be involved in the elevation of aminotransferases of HBeAg-negative patients, particularly those with low HBV-DNA levels. PMID:27124068

  14. Selected Cytokines Serve as Potential Biomarkers for Predicting Liver Inflammation and Fibrosis in Chronic Hepatitis B Patients With Normal to Mildly Elevated Aminotransferases.

    PubMed

    Deng, Yong-Qiong; Zhao, Hong; Ma, An-Lin; Zhou, Ji-Yuan; Xie, Shi-Bin; Zhang, Xu-Qing; Zhang, Da-Zhi; Xie, Qing; Zhang, Guo; Shang, Jia; Cheng, Jun; Zhao, Wei-Feng; Zou, Zhi-Qiang; Zhang, Ming-Xiang; Wang, Gui-Qiang

    2015-11-01

    Previous studies of small cohorts have implicated several circulating cytokines with progression of chronic hepatitis B (CHB). However, to date there have been no reliable biomarkers for assessing histological liver damage in CHB patients with normal or mildly elevated alanine aminotransferase (ALT). The aim of the present study was to investigate the association between circulating cytokines and histological liver damage in a large cohort. Also, this study was designed to assess the utility of circulating cytokines in diagnosing liver inflammation and fibrosis in CHB patients with ALT less than 2 times the upper limit of normal range (ULN). A total of 227 CHB patients were prospectively enrolled. All patients underwent liver biopsy and staging by Ishak system. Patients with at least moderate inflammation showed significantly higher levels of CXCL-11, CXCL-10, and interleukin (IL)-2 receptor (R) than patients with less than moderate inflammation (P < 0.001). Patients with significant fibrosis had higher levels of IL-8 (P = 0.027), transforming growth factor alpha (TGF-α) (P = 0.011), IL-2R (P = 0.002), and CXCL-11 (P = 0.032) than the group without significant fibrosis. In addition, 31.8% and 29.1% of 151 patients with ALT < 2 × ULN had at least moderate inflammation and significant fibrosis, respectively. Multivariate analysis demonstrated that CXCL-11 was independently associated with at least moderate inflammation, and TGF-α and IL-2R independently correlated with significant fibrosis in patients with ALT < 2 × ULN. Based on certain cytokines and clinical parameters, an inflammation-index and fib-index were developed, which showed areas under the receiver operating characteristics curve (AUROC) of 0.75 (95% CI 0.66-0.84) for at least moderate inflammation and 0.82 (95% CI 0.75-0.90) for significant fibrosis, correspondingly. Compared to existing scores, fib-index was significantly superior to aspartate aminotransferase

  15. Structural Analysis of WbpE from Pseudomonas aeruginosa PAO1: A Nucleotide Sugar Aminotransferase Involved in O-Antigen Assembly

    SciTech Connect

    Larkin, A.; Olivier, N; Imperiali, B

    2010-01-01

    In recent years, the opportunistic pathogen Pseudomonas aeruginosa has emerged as a major source of hospital-acquired infections. Effective treatment has proven increasingly difficult due to the spread of multidrug resistant strains and thus requires a deeper understanding of the biochemical mechanisms of pathogenicity. The central carbohydrate of the P. aeruginosa PAO1 (O5) B-band O-antigen, ManNAc(3NAc)A, has been shown to be critical for virulence and is produced in a stepwise manner by five enzymes in the Wbp pathway (WbpA, WbpB, WbpE, WbpD, and WbpI). Herein, we present the crystal structure of the aminotransferase WbpE from P. aeruginosa PAO1 in complex with the cofactor pyridoxal 5{prime}-phosphate (PLP) and product UDP-GlcNAc(3NH{sub 2})A as the external aldimine at 1.9 {angstrom} resolution. We also report the structures of WbpE in complex with PMP alone as well as the PLP internal aldimine and show that the dimeric structure of WbpE observed in the crystal structure is confirmed by analytical ultracentrifugation. Analysis of these structures reveals that the active site of the enzyme is composed of residues from both subunits. In particular, we show that a key residue (Arg229), which has previously been implicated in direct interactions with the {alpha}-carboxylate moiety of {alpha}-ketoglutarate, is also uniquely positioned to bestow specificity for the 6{double_prime}-carboxyl group of GlcNAc(3NH2)A through a salt bridge. This finding is intriguing because while an analogous basic residue is present in WbpE homologues that do not process 6{double_prime}-carboxyl-modified saccharides, recent structural studies reveal that this side chain is retracted to accommodate a neutral C6{double_prime} atom. This work represents the first structural analysis of a nucleotide sugar aminotransferase with a bound product modified at the C2{double_prime}, C3{double_prime}, and C6{double_prime} positions and provides insight into a novel target for treatment of P

  16. Selected Cytokines Serve as Potential Biomarkers for Predicting Liver Inflammation and Fibrosis in Chronic Hepatitis B Patients With Normal to Mildly Elevated Aminotransferases

    PubMed Central

    Deng, Yong-Qiong; Zhao, Hong; Ma, An-Lin; Zhou, Ji-Yuan; Xie, Shi-Bin; Zhang, Xu-Qing; Zhang, Da-Zhi; Xie, Qing; Zhang, Guo; Shang, Jia; Cheng, Jun; Zhao, Wei-Feng; Zou, Zhi-Qiang; Zhang, Ming-Xiang; Wang, Gui-Qiang

    2015-01-01

    Abstract Previous studies of small cohorts have implicated several circulating cytokines with progression of chronic hepatitis B (CHB). However, to date there have been no reliable biomarkers for assessing histological liver damage in CHB patients with normal or mildly elevated alanine aminotransferase (ALT). The aim of the present study was to investigate the association between circulating cytokines and histological liver damage in a large cohort. Also, this study was designed to assess the utility of circulating cytokines in diagnosing liver inflammation and fibrosis in CHB patients with ALT less than 2 times the upper limit of normal range (ULN). A total of 227 CHB patients were prospectively enrolled. All patients underwent liver biopsy and staging by Ishak system. Patients with at least moderate inflammation showed significantly higher levels of CXCL-11, CXCL-10, and interleukin (IL)-2 receptor (R) than patients with less than moderate inflammation (P < 0.001). Patients with significant fibrosis had higher levels of IL-8 (P = 0.027), transforming growth factor alpha (TGF-α) (P = 0.011), IL-2R (P = 0.002), and CXCL-11 (P = 0.032) than the group without significant fibrosis. In addition, 31.8% and 29.1% of 151 patients with ALT < 2 × ULN had at least moderate inflammation and significant fibrosis, respectively. Multivariate analysis demonstrated that CXCL-11 was independently associated with at least moderate inflammation, and TGF-α and IL-2R independently correlated with significant fibrosis in patients with ALT < 2 × ULN. Based on certain cytokines and clinical parameters, an inflammation-index and fib-index were developed, which showed areas under the receiver operating characteristics curve (AUROC) of 0.75 (95% CI 0.66–0.84) for at least moderate inflammation and 0.82 (95% CI 0.75–0.90) for significant fibrosis, correspondingly. Compared to existing scores, fib-index was significantly superior to aspartate

  17. Risk factors associated with hepatitis B or C markers or elevated alanine aminotransferase level among blood donors on a tropical island: the Guadeloupe experience.

    PubMed

    Fest, T; Viel, J F; Agis, F; Coffe, C; Dupond, J L; Hervé, P

    1992-10-01

    Donated blood is currently screened for hepatitis B surface antigen (HBsAg), antibody to hepatitis B core antigen (anti-HBc), antibody to hepatitis C virus (anti-HCV), and alanine aminotransferase (ALT) levels to prevent posttransfusion hepatitis. A prospective study of 2368 blood donors was carried out in Guadeloupe (French West Indies) with a view to determining the risk factors associated with serologic abnormalities. Blood donors included in the study had to complete a questionnaire. Statistical analysis was performed on the data thus obtained: 571 donations (24%) were positive for at least one of the four analyzed markers. The results were that 3.2 percent were positive for HBsAg, 22 percent for anti-HBc, and 0.8 percent for anti-HCV, and 1.4 percent had ALT > or = 45 IU per L. A good correlation was found between anti-HCV and elevated ALT. Transfusion history and two socioeconomic categories (working class, military personnel) were found to be risk factors. Other risk factors were lifelong residence in Guadeloupe (with risk increasing with the number of years), birthplace and current residence in the southern part of the island, and the existence of gastrointestinal discomfort unrelated to viral hepatitis (odds ratio = 2.98). The results of this study illustrate the difficulty of implementing a preventive policy against posttransfusion hepatitis in a tropical area. The unique epidemiologic situation of Guadeloupe as regards hepatitis B virus has led to more restrictive criteria for the acceptance of blood donors.

  18. vanishing tassel2 Encodes a Grass-Specific Tryptophan Aminotransferase Required for Vegetative and Reproductive Development in Maize[C][W][OA

    PubMed Central

    Phillips, Kimberly A.; Skirpan, Andrea L.; Liu, Xing; Christensen, Ashley; Slewinski, Thomas L.; Hudson, Christopher; Barazesh, Solmaz; Cohen, Jerry D.; Malcomber, Simon; McSteen, Paula

    2011-01-01

    Auxin plays a fundamental role in organogenesis in plants. Multiple pathways for auxin biosynthesis have been proposed, but none of the predicted pathways are completely understood. Here, we report the positional cloning and characterization of the vanishing tassel2 (vt2) gene of maize (Zea mays). Phylogenetic analyses indicate that vt2 is a co-ortholog of TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS1 (TAA1), which converts Trp to indole-3-pyruvic acid in one of four hypothesized Trp-dependent auxin biosynthesis pathways. Unlike single mutations in TAA1, which cause subtle morphological phenotypes in Arabidopsis thaliana, vt2 mutants have dramatic effects on vegetative and reproductive development. vt2 mutants share many similarities with sparse inflorescence1 (spi1) mutants in maize. spi1 is proposed to encode an enzyme in the tryptamine pathway for Trp-dependent auxin biosynthesis, although this biochemical activity has recently been questioned. Surprisingly, spi1 vt2 double mutants had only a slightly more severe phenotype than vt2 single mutants. Furthermore, both spi1 and vt2 single mutants exhibited a reduction in free auxin levels, but the spi1 vt2 double mutants did not have a further reduction compared with vt2 single mutants. Therefore, both spi1 and vt2 function in auxin biosynthesis in maize, possibly in the same pathway rather than independently as previously proposed. PMID:21335375

  19. The consensus-based approach for gene/enzyme replacement therapies and crystallization strategies: the case of human alanine-glyoxylate aminotransferase.

    PubMed

    Mesa-Torres, Noel; Yunta, Cristina; Fabelo-Rosa, Israel; Gonzalez-Rubio, Juana María; Sánchez-Ruiz, José M; Salido, Eduardo; Albert, Armando; Pey, Angel L

    2014-09-15

    Protein stability is a fundamental issue in biomedical and biotechnological applications of proteins. Among these applications, gene- and enzyme-replacement strategies are promising approaches to treat inherited diseases that may benefit from protein engineering techniques, even though these beneficial effects have been largely unexplored. In the present study we apply a sequence-alignment statistics procedure (consensus-based approach) to improve the activity and stability of the human AGT (alanine-glyoxylate aminotransferase) protein, an enzyme which causes PH1 (primary hyperoxaluria type I) upon mutation. By combining only five consensus mutations, we obtain a variant (AGT-RHEAM) with largely enhanced in vitro thermal and kinetic stability, increased activity, and with no side effects on foldability and peroxisomal targeting in mammalian cells. The structure of AGT-RHEAM reveals changes at the dimer interface and improved electrostatic interactions responsible for increased kinetic stability. Consensus-based variants maintained the overall protein fold, crystallized more easily and improved the expression as soluble proteins in two different systems [AGT and CIPK24 (CBL-interacting serine/threonine-protein kinase) SOS2 (salt-overly-sensitive 2)]. Thus the consensus-based approach also emerges as a simple and generic strategy to increase the crystallization success for hard-to-get protein targets as well as to enhance protein stability and function for biomedical applications.

  20. BRANCHED-CHAIN AMINOTRANSFERASE4 Is Part of the Chain Elongation Pathway in the Biosynthesis of Methionine-Derived Glucosinolates in Arabidopsis[W

    PubMed Central

    Schuster, Joachim; Knill, Tanja; Reichelt, Michael; Gershenzon, Jonathan; Binder, Stefan

    2006-01-01

    As part of our analysis of branched-chain amino acid metabolism in plants, we analyzed the function of Arabidopsis thaliana BRANCHED-CHAIN AMINOTRANSFERASE4 (BCAT4). Recombinant BCAT4 showed high efficiency with Met and its derivatives and the corresponding 2-oxo acids, suggesting its participation in the chain elongation pathway of Met-derived glucosinolate biosynthesis. This was substantiated by in vivo analysis of two BCAT4 T-DNA knockout mutants, in which Met-derived aliphatic glucosinolate accumulation is reduced by ∼50%. The increase in free Met and S-methylmethionine levels in these mutants, together with in vitro substrate specificity, strongly implicate BCAT4 in catalysis of the initial deamination of Met to 4-methylthio-2-oxobutyrate. BCAT4 transcription is induced by wounding and is predominantly observed in the phloem. BCAT4 transcript accumulation also follows a diurnal rhythm, and green fluorescent protein tagging experiments and subcellular protein fractions show that BCAT4 is located in the cytosol. The assignment of BCAT4 to the Met chain elongation pathway documents the close evolutionary relationship of this pathway to Leu biosynthesis. In addition to BCAT4, the enzyme methylthioalkylmalate synthase 1 has been recruited for the Met chain elongation pathway from a gene family involved in Leu formation. This suggests that the two pathways have a common evolutionary origin. PMID:17056707

  1. Biochemical and phylogenetic characterization of a novel diaminopimelate biosynthesis pathway in prokaryotes identifies a diverged form of LL-diaminopimelate aminotransferase.

    PubMed

    Hudson, André O; Gilvarg, Charles; Leustek, Thomas

    2008-05-01

    A variant of the diaminopimelate (DAP)-lysine biosynthesis pathway uses an LL-DAP aminotransferase (DapL, EC 2.6.1.83) to catalyze the direct conversion of L-2,3,4,5-tetrahydrodipicolinate to LL-DAP. Comparative genomic analysis and experimental verification of DapL candidates revealed the existence of two diverged forms of DapL (DapL1 and DapL2). DapL orthologs were identified in eubacteria and archaea. In some species the corresponding dapL gene was found to lie in genomic contiguity with other dap genes, suggestive of a polycistronic structure. The DapL candidate enzymes were found to cluster into two classes sharing approximately 30% amino acid identity. The function of selected enzymes from each class was studied. Both classes were able to functionally complement Escherichia coli dapD and dapE mutants and to catalyze LL-DAP transamination, providing functional evidence for a role in DAP/lysine biosynthesis. In all cases the occurrence of dapL in a species correlated with the absence of genes for dapD and dapE representing the acyl DAP pathway variants, and only in a few cases was dapL coincident with ddh encoding meso-DAP dehydrogenase. The results indicate that the DapL pathway is restricted to specific lineages of eubacteria including the Cyanobacteria, Desulfuromonadales, Firmicutes, Bacteroidetes, Chlamydiae, Spirochaeta, and Chloroflexi and two archaeal groups, the Methanobacteriaceae and Archaeoglobaceae.

  2. Gly161 mutations associated with Primary Hyperoxaluria Type I induce the cytosolic aggregation and the intracellular degradation of the apo-form of alanine:glyoxylate aminotransferase.

    PubMed

    Oppici, Elisa; Roncador, Alessandro; Montioli, Riccardo; Bianconi, Silvia; Cellini, Barbara

    2013-12-01

    Primary Hyperoxaluria Type I (PH1) is a severe rare disorder of metabolism due to inherited mutations on liver peroxisomal alanine:glyoxylate aminotransferase (AGT), a pyridoxal 5'-phosphate (PLP)-dependent enzyme whose deficiency causes the deposition of calcium oxalate crystals in the kidneys and urinary tract. PH1 is an extremely heterogeneous disease and there are more than 150 disease-causing mutations currently known, most of which are missense mutations. Moreover, the molecular mechanisms by which missense mutations lead to AGT deficiency span from structural, functional to subcellular localization defects. Gly161 is a highly conserved residue whose mutation to Arg, Cys or Ser is associated with PH1. Here we investigated the molecular bases of the AGT deficit caused by Gly161 mutations with expression studies in a mammalian cellular system paired with biochemical analyses on the purified recombinant proteins. Our results show that the mutations of Gly161 (i) strongly reduce the expression levels and the intracellular half-life of AGT, and (ii) make the protein in the apo-form prone to an electrostatically-driven aggregation in the cell cytosol. The coenzyme PLP, by shifting the equilibrium from the apo- to the holo-form, is able to reduce the aggregation propensity of the variants, thus partly decreasing the effect of the mutations. Altogether, these results shed light on the mechanistic details underlying the pathogenicity of Gly161 variants, thus expanding our knowledge of the enzymatic phenotypes leading to AGT deficiency.

  3. Glutamine-Glutamate Cycle Flux Is Similar in Cultured Astrocytes and Brain and Both Glutamate Production and Oxidation Are Mainly Catalyzed by Aspartate Aminotransferase.

    PubMed

    Hertz, Leif; Rothman, Douglas L

    2017-02-24

    The glutamine-glutamate cycle provides neurons with astrocyte-generated glutamate/γ-aminobutyric acid (GABA) and oxidizes glutamate in astrocytes, and it returns released transmitter glutamate/GABA to neurons after astrocytic uptake. This review deals primarily with the glutamate/GABA generation/oxidation, although it also shows similarity between metabolic rates in cultured astrocytes and intact brain. A key point is identification of the enzyme(s) converting astrocytic α-ketoglutarate to glutamate and vice versa. Most experiments in cultured astrocytes, including those by one of us, suggest that glutamate formation is catalyzed by aspartate aminotransferase (AAT) and its degradation by glutamate dehydrogenase (GDH). Strongly supported by results shown in Table 1 we now propose that both reactions are primarily catalyzed by AAT. This is possible because the formation occurs in the cytosol and the degradation in mitochondria and they are temporally separate. High glutamate/glutamine concentrations abolish the need for glutamate production from α-ketoglutarate and due to metabolic coupling between glutamate synthesis and oxidation these high concentrations render AAT-mediated glutamate oxidation impossible. This necessitates the use of GDH under these conditions, shown by insensitivity of the oxidation to the transamination inhibitor aminooxyacetic acid (AOAA). Experiments using lower glutamate/glutamine concentration show inhibition of glutamate oxidation by AOAA, consistent with the coupled transamination reactions described here.

  4. Novel loss-of-function putative aminotransferase alleles cause biosynthesis of capsinoids, nonpungent capsaicinoid analogues, in mildly pungent chili peppers (Capsicum chinense).

    PubMed

    Tanaka, Yoshiyuki; Hosokawa, Munetaka; Miwa, Tetsuya; Watanabe, Tatsuo; Yazawa, Susumu

    2010-11-24

    Capsinoids are a group of nonpungent capsaicinoid analogues produced in Capsicum fruits. They have similar bioactivities to capsaicinoids such as suppression of fat accumulation and antioxidant activity. They are more palatable ingredients in dietary supplements than capsaicinoids because of their low pungency. Previous studies on nonpungent Capsicum annuum cultivars showed that capsinoid biosynthesis is caused by loss-of-function putative aminotransferase (p-amt) alleles. This study showed that three mildly pungent cultivars of Capsicum chinense (Zavory Hot, Aji Dulce strain 2, and Belize Sweet) contain high levels of capsinoid. It was shown that these cultivars have novel p-amt alleles, which contain mutations that differ from those of C. annuum. Sequence analysis of p-amt in Belize Sweet revealed that a 5 bp insertion (TGGGC) results in a frameshift mutation. A transposable element (Tcc) was found in the p-amt of Zavory Hot and Aji Dulce strain 2. Tcc has features similar to those of the hAT transposon family. This was inserted in the fifth intron of Zavory Hot and in third intron of Aji Dulce strain 2. The p-amt alleles harboring Tcc cannot produce an active p-AMT. These mildly pungent cultivars will provide a new natural source of capsinoids.

  5. Complete amino acid sequence of branched-chain amino acid aminotransferase (transaminase B) of Salmonella typhimurium, identification of the coenzyme-binding site and sequence comparison analysis

    SciTech Connect

    Feild, M.J.

    1988-01-01

    The complete amino acid sequence of the subunit of branched-chain amino acid aminotransferase of Salmonella typhimurium was determined by automated Edman degradation of peptide fragments generated by chemical and enzymatic digestion of S-carboxymethylated and S-pyridylethylated transaminase B. Peptide fragments of transaminase B were generated by treatment of the enzyme with trypsin, Staphylococcus aureus V8 protease, endoproteinase Lys-C, and cyanogen bromide. Protocols were developed for separation of the peptide fragments by reverse-phase high performance liquid chromatography (HPLC), ion-exchange HPLC, and SDS-urea gel electrophoresis. The enzyme subunit contains 308 amino acid residues and has a molecular weight of 33,920 daltons. The coenzyme-binding site was determined by treatment of the enzyme, containing bound pyridoxal 5-phosphate, with tritiated sodium borohydride prior to trypsin digestion. Monitoring radioactivity incorporation and peptide map comparisons with an apoenzyme tryptic digest, allowed identification of the pyridoxylated-peptide which was isolated by reverse-phase HPLC and sequenced. The coenzyme-binding site is a lysyl residue at position 159. Some peptides were further characterized by fast atom bombardment mass spectrometry.

  6. Glutamine-Glutamate Cycle Flux Is Similar in Cultured Astrocytes and Brain and Both Glutamate Production and Oxidation Are Mainly Catalyzed by Aspartate Aminotransferase

    PubMed Central

    Hertz, Leif; Rothman, Douglas L

    2017-01-01

    The glutamine-glutamate cycle provides neurons with astrocyte-generated glutamate/γ-aminobutyric acid (GABA) and oxidizes glutamate in astrocytes, and it returns released transmitter glutamate/GABA to neurons after astrocytic uptake. This review deals primarily with the glutamate/GABA generation/oxidation, although it also shows similarity between metabolic rates in cultured astrocytes and intact brain. A key point is identification of the enzyme(s) converting astrocytic α-ketoglutarate to glutamate and vice versa. Most experiments in cultured astrocytes, including those by one of us, suggest that glutamate formation is catalyzed by aspartate aminotransferase (AAT) and its degradation by glutamate dehydrogenase (GDH). Strongly supported by results shown in Table 1 we now propose that both reactions are primarily catalyzed by AAT. This is possible because the formation occurs in the cytosol and the degradation in mitochondria and they are temporally separate. High glutamate/glutamine concentrations abolish the need for glutamate production from α-ketoglutarate and due to metabolic coupling between glutamate synthesis and oxidation these high concentrations render AAT-mediated glutamate oxidation impossible. This necessitates the use of GDH under these conditions, shown by insensitivity of the oxidation to the transamination inhibitor aminooxyacetic acid (AOAA). Experiments using lower glutamate/glutamine concentration show inhibition of glutamate oxidation by AOAA, consistent with the coupled transamination reactions described here. PMID:28245547

  7. Dual roles of a conserved pair, Arg23 and Ser20, in recognition of multiple substrates in {alpha}-aminoadipate aminotransferase from Thermus thermophilus

    SciTech Connect

    Ouchi, Takuya; Tomita, Takeo; Miyagawa, Tomoharu; Kuzuyama, Tomohisa; Nishiyama, Makoto

    2009-10-09

    To clarify the mechanism for substrate recognition of {alpha}-aminoadipate aminotransferase (AAA-AT) from Thermus thermophilus, the crystal structure of AAA-AT complexed with N-(5'-phosphopyridoxyl)-L-glutamate (PPE) was determined at 1.67 A resolution. The crystal structure revealed that PPE is recognized by amino acid residues the same as those seen in N-(5'-phosphopyridoxyl)-L-{alpha}-aminoadipate (PPA) recognition; however, to bind the {gamma}-carboxyl group of Glu at a fixed position, the C{alpha} atom of the Glu moiety moves 0.80 A toward the {gamma}-carboxyl group in the PPE complex. Markedly decreased activity for Asp can be explained by the shortness of the aspartyl side chain to be recognized by Arg23 and further dislocation of the C{alpha} atom of bound Asp. Site-directed mutagenesis revealed that Arg23 has dual functions for reaction, (i) recognition of {gamma} ({delta})-carboxyl group of Glu (AAA) and (ii) rearrangement of {alpha}2 helix by changing the interacting partners to place the hydrophobic substrate at the suitable position.

  8. Diabetes-linked transcription factor HNF4α regulates metabolism of endogenous methylarginines and β-aminoisobutyric acid by controlling expression of alanine-glyoxylate aminotransferase 2

    PubMed Central

    Burdin, Dmitry V.; Kolobov, Alexey A.; Brocker, Chad; Soshnev, Alexey A.; Samusik, Nikolay; Demyanov, Anton V.; Brilloff, Silke; Jarzebska, Natalia; Martens-Lobenhoffer, Jens; Mieth, Maren; Maas, Renke; Bornstein, Stefan R.; Bode-Böger, Stefanie M.; Gonzalez, Frank; Weiss, Norbert; Rodionov, Roman N.

    2016-01-01

    Elevated levels of circulating asymmetric and symmetric dimethylarginines (ADMA and SDMA) predict and potentially contribute to end organ damage in cardiovascular diseases. Alanine-glyoxylate aminotransferase 2 (AGXT2) regulates systemic levels of ADMA and SDMA, and also of beta-aminoisobutyric acid (BAIB)-a modulator of lipid metabolism. We identified a putative binding site for hepatic nuclear factor 4 α (HNF4α) in AGXT2 promoter sequence. In a luciferase reporter assay we found a 75% decrease in activity of Agxt2 core promoter after disruption of the HNF4α binding site. Direct binding of HNF4α to Agxt2 promoter was confirmed by chromatin immunoprecipitation assay. siRNA-mediated knockdown of Hnf4a led to an almost 50% reduction in Agxt2 mRNA levels in Hepa 1–6 cells. Liver-specific Hnf4a knockout mice exhibited a 90% decrease in liver Agxt2 expression and activity, and elevated plasma levels of ADMA, SDMA and BAIB, compared to wild-type littermates. Thus we identified HNF4α as a major regulator of Agxt2 expression. Considering a strong association between human HNF4A polymorphisms and increased risk of type 2 diabetes our current findings suggest that downregulation of AGXT2 and subsequent impairment in metabolism of dimethylarginines and BAIB caused by HNF4α deficiency might contribute to development of cardiovascular complications in diabetic patients. PMID:27752141

  9. Diabetes-linked transcription factor HNF4α regulates metabolism of endogenous methylarginines and β-aminoisobutyric acid by controlling expression of alanine-glyoxylate aminotransferase 2.

    PubMed

    Burdin, Dmitry V; Kolobov, Alexey A; Brocker, Chad; Soshnev, Alexey A; Samusik, Nikolay; Demyanov, Anton V; Brilloff, Silke; Jarzebska, Natalia; Martens-Lobenhoffer, Jens; Mieth, Maren; Maas, Renke; Bornstein, Stefan R; Bode-Böger, Stefanie M; Gonzalez, Frank; Weiss, Norbert; Rodionov, Roman N

    2016-10-18

    Elevated levels of circulating asymmetric and symmetric dimethylarginines (ADMA and SDMA) predict and potentially contribute to end organ damage in cardiovascular diseases. Alanine-glyoxylate aminotransferase 2 (AGXT2) regulates systemic levels of ADMA and SDMA, and also of beta-aminoisobutyric acid (BAIB)-a modulator of lipid metabolism. We identified a putative binding site for hepatic nuclear factor 4 α (HNF4α) in AGXT2 promoter sequence. In a luciferase reporter assay we found a 75% decrease in activity of Agxt2 core promoter after disruption of the HNF4α binding site. Direct binding of HNF4α to Agxt2 promoter was confirmed by chromatin immunoprecipitation assay. siRNA-mediated knockdown of Hnf4a led to an almost 50% reduction in Agxt2 mRNA levels in Hepa 1-6 cells. Liver-specific Hnf4a knockout mice exhibited a 90% decrease in liver Agxt2 expression and activity, and elevated plasma levels of ADMA, SDMA and BAIB, compared to wild-type littermates. Thus we identified HNF4α as a major regulator of Agxt2 expression. Considering a strong association between human HNF4A polymorphisms and increased risk of type 2 diabetes our current findings suggest that downregulation of AGXT2 and subsequent impairment in metabolism of dimethylarginines and BAIB caused by HNF4α deficiency might contribute to development of cardiovascular complications in diabetic patients.

  10. L-alanine-glyoxylate aminotransferase II of rat kidney and liver mitochondria possesses cysteine S-conjugate beta-lyase activity: a contributing factor to the nephrotoxicity/hepatotoxicity of halogenated alkenes?

    PubMed Central

    Cooper, Arthur J L; Krasnikov, Boris F; Okuno, Etsuo; Jeitner, Thomas M

    2003-01-01

    Several halogenated alkenes are metabolized in part to cysteine S-conjugates, which are mitochondrial toxicants of kidney and, to a lesser extent, other organs. Toxicity is due to cysteine S-conjugate beta-lyases, which convert the cysteine S-conjugate into pyruvate, ammonia and a reactive sulphur-containing fragment. A section of the human population is exposed to halogenated alkenes. To understand the health effects of such exposure, it is important to identify cysteine S-conjugate beta-lyases that contribute to mitochondrial damage. Mitochondrial aspartate aminotransferase [Cooper, Bruschi, Iriarte and Martinez-Carrion (2002) Biochem. J. 368, 253-261] and mitochondrial branched-chain aminotransferase [Cooper, Bruschi, Conway and Hutson (2003) Biochem. Pharmacol. 65, 181-192] exhibit beta-lyase activity toward S -(1,2-dichlorovinyl)-L-cysteine (the cysteine S-conjugate of trichloroethylene) and S -(1,1,2,2-tetrafluoroethyl)-L-cysteine (the cysteine S-conjugate of tetrafluoroethylene). Turnover leads to eventual inactivation of these enzymes. Here we report that mitochondrial L-alanine-glyoxylate aminotransferase II, which, in the rat, is most active in kidney, catalyses cysteine S-conjugate beta-lyase reactions with S -(1,1,2,2-tetrafluoroethyl)-L-cysteine, S -(1,2-dichlorovinyl)-L-cysteine and S -(benzothiazolyl-L-cysteine); turnover leads to inactivation. Previous workers showed that the reactive-sulphur-containing fragment released from S -(1,1,2,2-tetrafluoroethyl)-L-cysteine and S -(1,2-dichlorovinyl)-L-cysteine is toxic by acting as a thioacylating agent - particularly of lysine residues in nearby proteins. Toxicity, however, may also involve 'self-inactivation' of key enzymes. The present findings suggest that alanine-glyoxylate aminotransferase II may be an important factor in the well-established targeting of rat kidney mitochondria by toxic halogenated cysteine S-conjugates. Previous reports suggest that alanine-glyoxylate aminotransferase II is absent

  11. A gene encoding lysine 6-aminotransferase, which forms the beta-lactam precursor alpha-aminoadipic acid, is located in the cluster of cephamycin biosynthetic genes in Nocardia lactamdurans.

    PubMed Central

    Coque, J J; Liras, P; Laiz, L; Martín, J F

    1991-01-01

    A gene (lat) encoding lysine 6-aminotransferase was found upstream of the pcbAB (encoding alpha-aminoadipylcysteinyl-valine synthetase) and pcbC (encoding isopenicillin N synthase) genes in the cluster of early cephamycin biosynthetic genes in Nocardia lactamdurans. The lat gene was separated by a small intergenic region of 64 bp from the 5' end of the pcbAB gene. The lat gene contained an open reading frame of 1,353 nucleotides (71.4% G + C) encoding a protein of 450 amino acids with a deduced molecular mass of 48,811 Da. Expression of DNA fragments carrying the lat gene in Streptomyces lividans led to a high lysine 6-aminotransferase activity which was absent from untransformed S. lividans. The enzyme was partially purified from S. lividans(pULBS8) and showed a molecular mass of 52,800 Da as calculated by Sephadex gel filtration and polyacrylamide gel electrophoresis. DNA sequences which hybridized strongly with the lat gene of N. lactamdurans were found in four cephamycin-producing Streptomyces species but not in four other actinomycetes which are not known to produce beta-lactams, suggesting that the gene is specific for beta-lactam biosynthesis and is not involved in general lysine catabolism. The protein encoded by the lat gene showed similarity to ornithine-5-aminotransferases and N-acetylornithine-5-aminotransferases and contained a pyridoxal phosphate-binding consensus amino acid sequence around Lys-300 of the protein. The evolutionary implications of the lat gene as a true beta-lactam biosynthetic gene are discussed. Images PMID:1917857

  12. The ribavirin analog ICN 17261 demonstrates reduced toxicity and antiviral effects with retention of both immunomodulatory activity and reduction of hepatitis-induced serum alanine aminotransferase levels.

    PubMed

    Tam, R C; Ramasamy, K; Bard, J; Pai, B; Lim, C; Averett, D R

    2000-05-01

    The demonstrated utility of the nucleoside analog ribavirin in the treatment of certain viral diseases can be ascribed to its multiple distinct properties. These properties may vary in relative importance in differing viral disease conditions and include the direct inhibition of viral replication, the promotion of T-cell-mediated immune responses via an enhanced type 1 cytokine response, and a reduction of circulating alanine aminotransferase (ALT) levels associated with hepatic injury. Ribavirin also has certain known toxicities, including the induction of anemia upon chronic administration. To determine if all these properties are linked, we compared the D-nucleoside ribavirin to its L-enantiomer (ICN 17261) with regard to these properties. Strong similarities were seen for these two compounds with respect to induction of type 1 cytokine bias in vitro, enhancement of type 1 cytokine responses in vivo, and the reduction of serum ALT levels in a murine hepatitis model. In contrast, ICN 17261 had no in vitro antiviral activity against a panel of RNA and DNA viruses, while ribavirin exhibited its characteristic activity profile. Importantly, the preliminary in vivo toxicology profile of ICN 17261 is significantly more favorable than that of ribavirin. Administration of 180 mg of ICN 17261 per kg of body weight to rats by oral gavage for 4 weeks generated substantial serum levels of drug but no observable clinical pathology, whereas equivalent doses of ribavirin induced a significant anemia and leukopenia. Thus, structural modification of ribavirin can dissociate its immunomodulatory properties from its antiviral and toxicologic properties, resulting in a compound (ICN 17261) with interesting therapeutic potential.

  13. Contributions of the substrate-binding arginine residues to maleate-induced closure of the active site of Escherichia coli aspartate aminotransferase.

    PubMed

    Matharu, A; Hayashi, H; Kagamiyama, H; Maras, B; John, R A

    2001-03-01

    Crystallography shows that aspartate aminotransferase binds dicarboxylate substrate analogues by bonds to Arg292 and Arg386, respectively [Jager, J, Moser, M. Sauder, U. & Jansonius, J. N. (1994) J. Mol. Biol., 239, 285-305]. The contribution of each interaction to the conformational change that the enzyme undergoes when it binds ligands via these residues, is assessed by probing mutant forms of the enzyme lacking either or both arginines. The probes used are NaH(3)BCN which reduces the cofactor imine, the reactive substrate analogue, cysteine sulfinate and proteolysis by trypsin. The unreactive substrate analogue, maleate, is used to induce closure. Each single mutant reacted only 2.5-fold more slowly with NaH(3)BCN than the wild-type indicating that charge repulsion by the arginines contributes little to maintaining the open conformation. Maleate lowered the rate of reduction of the wild-type enzyme more than 300-fold but had little effect on the reaction of the mutant enzymes indicating that the ability of this dicarboxylate analogue to bridge the arginines precisely makes the major contribution to closure. The R292L mutant reacted 20 times more rapidly with cysteine sulfinate than R386L but 5 x 10(4) times more slowly than the wild-type enzyme, consistent with the proposal that enzyme's catalytic abilities are not developed unless closure is induced by bridging of the arginines. Proteolysis of the mutants with trypsin showed that, in the wild-type enzyme, the bonds most susceptible to trypsin are those contributed by Arg292 and Arg386. Proteolysis of the next most susceptible bond, at Arg25 in the double mutant, was protected by maleate demonstrating the presence of an additional site on the enzyme for binding dicarboxylates.

  14. Misfolding caused by the pathogenic mutation G47R on the minor allele of alanine:glyoxylate aminotransferase and chaperoning activity of pyridoxine.

    PubMed

    Montioli, Riccardo; Oppici, Elisa; Dindo, Mirco; Roncador, Alessandro; Gotte, Giovanni; Cellini, Barbara; Borri Voltattorni, Carla

    2015-10-01

    Liver peroxisomal alanine:glyoxylate aminotransferase (AGT), a pyridoxal 5'-phosphate (PLP) enzyme, exists as two polymorphic forms, the major (AGT-Ma) and the minor (AGT-Mi) haplotype. Deficit of AGT causes Primary Hyperoxaluria Type 1 (PH1), an autosomal recessive rare disease. Although ~one-third of the 79 disease-causing missense mutations segregates on AGT-Mi, only few of them are well characterized. Here for the first time the molecular and cellular defects of G47R-Mi are reported. When expressed in Escherichia coli, the recombinant purified G47R-Mi variant exhibits only a 2.5-fold reduction of its kcat, and its apo form displays a remarkably decreased PLP binding affinity, increased dimer-monomer equilibrium dissociation constant value, susceptibility to thermal denaturation and to N-terminal region proteolytic cleavage, and aggregation propensity. When stably expressed in a mammalian cell line, we found ~95% of the intact form of the variant in the insoluble fraction, and proteolyzed (within the N-terminal region) and aggregated forms both in the soluble and insoluble fractions. Moreover, the intact and nicked forms have a peroxisomal and a mitochondrial localization, respectively. Unlike what already seen for G41R-Mi, exposure of G47R-Mi expressing cells to pyridoxine (PN) remarkably increases the expression level and the specific activity in a dose-dependent manner, reroutes all the protein to peroxisomes, and rescues its functionality. Although the mechanism of the different effect of PN on the variants G47R-Mi and G41R-Mi remains elusive, the chaperoning activity of PN may be of value in the therapy of patients bearing the G47R mutation.

  15. Enzymological and mutational analysis of a complex primary hyperoxaluria type I phenotype involving alanine: Glyoxylate aminotransferase peroxisome-to-mitochondrion mistargeting and intraperoxisomal aggregation

    SciTech Connect

    Danpure, C.J.; Purdue, P.E.; Allsop, J.; Lumb, M.J.; Jennings, P.R. ); Scheinman, J.I. ); Mauer, S.M. ); Davidson, N.O. )

    1993-08-01

    Primary hyperoxaluri type 1 (PH1) is a rare autosomal recessive disease caused by a deficiency of the liver-specific peroxisomal enzyme alanine:glyoxylate aminotransferase (AGT). Three unrelated PH1 patients, who possess a novel complex phenotype, are described. At the enzymological level, this phenotype is characterized by a complete, or nearly complete, absence of AGT catalytic activity and reduced AGT immunoreactivity. Unlike normal individuals in whom the AGT is confined to the peroxisomal matrix, the immunoreactive AGT in these three patients was distributed approximately equally between the peroxisomes and mitochondria. The peroxisomal AGT appeared to be aggregated into amorphous core-like structures in which no other peroxisomal enzymes could be identified. Mutational analysis of the AGT gene showed that two of the three patients were compound heterozygotes for two previously unrecognized point mutations which caused Gly41[yields]Arg and Phe152[yields]Iso amino acid substitutions. The third patient was shown to be a compound heterozygote for the Gly41[yields]Arg mutation and a previously recognized Gly170[yields]Arg mutation. All three patients were homozygous for the Pro11[yields]Leu polymorphism that had been found previously with a high allelic frequency in normal populations. It is suggested the the Phe152[yields]Iso and Gly170[yields]Arg substitutions, which are only eighteen residues apart and located in the same highly conserved internal region of 58 amino acids, might be involved in the inhibition of peroxisomal targeting and/or import of AGT and, in combination with the Pro11[yields]Leu polymorphism, be responsible for its aberrant mitochondrial compartmentalization. On the other hand, the Gly41[yields]Arg substitution, either in combination with the Pro11[yields]Leu polymorphism or by itself, is predicted to be responsible for the intraperoxisomal aggregation of the AGT protein. 50 refs., 8 figs., 4 tabs.

  16. Targeted disruption of murine ornithine aminotransferase gene: Surprising neonatal lethality rescued by arginine therapy and possible mouse model for gyrate atrophy

    SciTech Connect

    Wang, T.; Steel, G.; Valle, D.

    1994-09-01

    Deficiency of ornithine-{delta}-aminotransferase (OAT) in humans results in gyrate atrophy (GA), an autosomal recessive blinding disorder characterized by progressive chorioretinal degeneration and hyperornithinemia. GA patients are otherwise asymptomatic. The explanation for the unique susceptibility of retina to this inborn error is not known. Poor understanding of the pathophysiology hampers development of effective therapy. To address these problems, we undertook targeted disruption of the murine OAT gene. We isolated a 13 kb genomic DNA fragment containing the first five exons of OAT from an AB1 mini-genomic library and made a replacement type targeting vector with the NEOR gene inserted into OAT exon 3 at codon 40. Following electroporation and selection, 4.4% of G418 resistant ES clones had undergone homologous recombination. Chimeric animals produced by blastocyst injection transmitted the disrupted OAT allele. Mice homozygous for OAT disruption (-/-) have no detectable OAT activity in their tissues. Although normal at birth, they die within 10 to 48 hrs. We hypothesize this lethality results from a block in gut arginine biosynthesis, a pathway that requires {open_quotes}reverse{close_quotes} OAT flux and is apparently critical for rapidly growing neonatal mice. Amino acid measurements reveal extreme reduction in urea cycle intermediates in dying mice. With i.p. arginine administration for the first 14 days of life, we have successfully obtained adult (-/-) mice. Plasma amino acids on these mice show a 10-15 fold increase in ornithine and a 2 fold decrease in lysine, biochemical hallmarks of GA. ERG and retinal histopathologic studies are in progress to examine the ocular phenotypes. These mice will provide an invaluable model to test strategies to reduce ornithine accumulation and should also prove useful in understanding the pathophysiology of retina degeneration.

  17. Molecular and Functional Analyses Support a Role of Ornithine-δ-Aminotransferase in the Provision of Glutamate for Glutamine Biosynthesis during Pine Germination1[W

    PubMed Central

    Cañas, Rafael A.; Villalobos, David P.; Díaz-Moreno, Sara M.; Cánovas, Francisco M.; Cantón, Francisco R.

    2008-01-01

    We report the molecular characterization and functional analysis of a gene (PsδOAT) from Scots pine (Pinus sylvestris) encoding Orn-δ-aminotransferase (δ-OAT; EC 2.6.1.13), an enzyme of arginine metabolism. The deduced amino acid sequence contains a putative N-terminal signal peptide for mitochondrial targeting. The polypeptide is similar to other δ-OATs from plants, yeast, and mammals and encoded by a single-copy gene in pine. PsδOAT encodes a functional δ-OAT as determined by expression of the recombinant protein in Escherichia coli and analysis of the active enzyme. The expression of PsδOAT was undetectable in the embryo, but highly induced at early stages of germination and seedling development in all different organs. Transcript levels decreased in later developmental stages, although an increase was observed in lignified stems of 90-d-old plants. An increase of δ-OAT activity was observed in germinating embryos and seedlings and appears to mirror the observed alterations in PsδOAT transcript levels. Similar expression patterns were also observed for genes encoding arginase and isocitrate dehydrogenase. Transcripts of PsδOAT and the arginase gene were found widely distributed in different cell types of pine organs. Consistent with these results a metabolic pathway is proposed for the nitrogen flow from the megagametophyte to the developing seedling, which is also supported by the relative abundance of free amino acids in embryos and seedlings. Taken together, our data support that δ-OAT plays an important role in this process providing glutamate for glutamine biosynthesis during early pine growth. PMID:18621980

  18. Mechanism of Inactivation of γ-Aminobutyric Acid Aminotransferase by (1S ,3S)-3-Amino-4-difluoromethylene-1-cyclopentanoic Acid (CPP-115)

    DOE PAGES

    Lee, Hyunbeom; Doud, Emma H.; Wu, Rui; ...

    2015-01-23

    γ-Aminobutyric acid aminotransferase (GABA-AT) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that degrades GABA, the principal inhibitory neurotransmitter in mammalian cells. When the concentration of GABA falls below a threshold level, convulsions can occur. Inhibition of GABA-AT raises GABA levels in the brain, which can terminate seizures as well as have potential therapeutic applications in treating other neurological disorders, including drug addiction. Among the analogues that we previously developed, (1S,3S)-3-amino-4-difluoromethylene-1-cyclopentanoic acid (CPP-115) showed 187 times greater potency than that of vigabatrin, a known inactivator of GABA-AT and approved drug (Sabril) for the treatment of infantile spasms and refractory adult epilepsy. Recently,more » CPP-115 was shown to have no adverse effects in a Phase I clinical trial. Here we report a novel inactivation mechanism for CPP-115, a mechanism-based inactivator that undergoes GABA-AT-catalyzed hydrolysis of the difluoromethylene group to a carboxylic acid with concomitant loss of two fluoride ions and coenzyme conversion to pyridoxamine 5'-phosphate (PMP). The partition ratio for CPP-115 with GABA-AT is about 2000, releasing cyclopentanone-2,4-dicarboxylate (22) and two other precursors of this compound (20 and 21). Time-dependent inactivation occurs by a conformational change induced by the formation of the aldimine of 4-aminocyclopentane-1,3-dicarboxylic acid and PMP (20), which disrupts an electrostatic interaction between Glu270 and Arg445 to form an electrostatic interaction between Arg445 and the newly formed carboxylate produced by hydrolysis of the difluoromethylene group in CPP-115, resulting in a noncovalent, tightly bound complex. Ultimately, this represents a novel mechanism for inactivation of GABA-AT and a new approach for the design of mechanism-based inactivators in general.« less

  19. Analysis of alanine aminotransferase in various organs of soybean (Glycine max) and in dependence of different nitrogen fertilisers during hypoxic stress.

    PubMed

    Rocha, Marcio; Sodek, Ladaslav; Licausi, Francesco; Hameed, Muhammad Waqar; Dornelas, Marcelo Carnier; van Dongen, Joost T

    2010-10-01

    Alanine aminotransferase (AlaAT) catalyses the reversible conversion of pyruvate and glutamate into alanine and oxoglutarate. In soybean, two subclasses were identified, each represented by two highly similar members. To investigate the role of AlaAT during hypoxic stress in soybean, changes in transcript level of both subclasses were analysed together with the enzyme activity and alanine content of the tissue. Moreover, the dependency of AlaAT activity and gene expression was investigated in relation to the source of nitrogen supplied to the plants. Using semi-quantitative PCR, GmAlaAT genes were determined to be highest expressed in roots and nodules. Under normal growth conditions, enzyme activity of AlaAT was detected in all organs tested, with lowest activity in the roots. Upon waterlogging-induced hypoxia, AlaAT activity increased strongly. Concomitantly, alanine accumulated. During re-oxygenation, AlaAT activity remained high, but the transcript level and the alanine content decreased. Our results show a role for AlaAT in the catabolism of alanine during the initial period of re-oxygenation following hypoxia. GmAlaAT also responded to nitrogen availability in the solution during waterlogging. Ammonium as nitrogen source induced both gene expression and enzyme activity of AlaAT more than when nitrate was supplied in the nutrient solution. The work presented here indicates that AlaAT might not only be important during hypoxia, but also during the recovery phase after waterlogging, when oxygen is available to the tissue again.

  20. Quantitative structure-activity relationship and molecular docking studies of a series of quinazolinonyl analogues as inhibitors of gamma amino butyric acid aminotransferase.

    PubMed

    Abdulfatai, Usman; Uzairu, Adamu; Uba, Sani

    2017-01-01

    Quantitative structure-activity relationship and molecular docking studies were carried out on a series of quinazolinonyl analogues as anticonvulsant inhibitors. Density Functional Theory (DFT) quantum chemical calculation method was used to find the optimized geometry of the anticonvulsants inhibitors. Four types of molecular descriptors were used to derive a quantitative relation between anticonvulsant activity and structural properties. The relevant molecular descriptors were selected by Genetic Function Algorithm (GFA). The best model was validated and found to be statistically significant with squared correlation coefficient (R(2)) of 0.934, adjusted squared correlation coefficient (R(2)adj) value of 0.912, Leave one out (LOO) cross validation coefficient (Q(2)) value of 0.8695 and the external validation (R(2)pred) of 0.72. Docking analysis revealed that the best compound with the docking scores of -9.5 kcal/mol formed hydrophobic interaction and H-bonding with amino acid residues of gamma aminobutyric acid aminotransferase (GABAAT). This research has shown that the binding affinity generated was found to be better than the commercially sold anti-epilepsy drug, vigabatrin. Also, it was found to be better than the one reported by other researcher. Our QSAR model and molecular docking results corroborate with each other and propose the directions for the design of new inhibitors with better activity against GABAAT. The present study will help in rational drug design and synthesis of new selective GABAAT inhibitors with predetermined affinity and activity and provides valuable information for the understanding of interactions between GABAAT and the anticonvulsants inhibitors.

  1. Comparison of FIB-4 index and aspartate aminotransferase to platelet ratio index on carcinogenesis in chronic hepatitis B treated with entecavir

    PubMed Central

    Nishikawa, Hiroki; Nishijima, Norihiro; Enomoto, Hirayuki; Sakamoto, Azusa; Nasu, Akihiro; Komekado, Hideyuki; Nishimura, Takashi; Kita, Ryuichi; Kimura, Toru; Iijima, Hiroko; Nishiguchi, Shuhei; Osaki, Yukio

    2017-01-01

    Aims: We sought to compare the effects of FIB-4 index and aspartate aminotransferase to platelet ratio index (APRI) on hepatocellular carcinoma (HCC) incidence in chronic hepatitis B (CHB) patients undergoing entecavir (ETV) therapy. Patient and methods: A total of 338 nucleosides analogue therapy naïve CHB patients initially treated with ETV were analyzed. The optimal cutoff points in each continuous variable were determined by receiver operating curve (ROC) analysis. The effects of FIB-4 index and APRI on HCC incidence were compared using time-dependent ROC analysis and factors linked to HCC incidence were also examined using univariate and multivariate analyses. Results: There were 215 males and 123 females with the median age of 52 years and the median baseline HBV-DNA level of 6.6 log copies/ml. The median follow-up interval after the initiation of ETV therapy was 4.99 years. During the follow-up period, 33 patients (9.8%) developed HCC. The 3-, 5- 7-year cumulative HCC incidence rates in all cases were 4.4%, 9.2% and 13.5%, respectively. In the multivariate analysis, FIB-4 index revealed to be an independent predictor associated with HCC incidence, while APRI was not. In the time-dependent ROC analyses for all cases and for all subgroups analyses stratified by viral status or cirrhosis status, all area under the ROCs in each time point (2-, 3-, 4-, 5-, 6-, and 7-year) of FIB-4 index were higher than those of APRI. Conclusion: FIB-4 index rather than APRI can be a useful predictor associated with HCC development for CHB patients undergoing ETV therapy. PMID:28243319

  2. Crystal structure of the S187F variant of human liver alanine: glyoxylate [corrected] aminotransferase associated with primary hyperoxaluria type I and its functional implications.

    PubMed

    Oppici, Elisa; Fodor, Krisztian; Paiardini, Alessandro; Williams, Chris; Voltattorni, Carla Borri; Wilmanns, Matthias; Cellini, Barbara

    2013-08-01

    The substitution of Ser187, a residue located far from the active site of human liver peroxisomal alanine:glyoxylate aminotransferase (AGT), by Phe gives rise to a variant associated with primary hyperoxaluria type I. Unexpectedly, previous studies revealed that the recombinant form of S187F exhibits a remarkable loss of catalytic activity, an increased pyridoxal 5'-phosphate (PLP) binding affinity and a different coenzyme binding mode compared with normal AGT. To shed light on the structural elements responsible for these defects, we solved the crystal structure of the variant to a resolution of 2.9 Å. Although the overall conformation of the variant is similar to that of normal AGT, we noticed: (i) a displacement of the PLP-binding Lys209 and Val185, located on the re and si side of PLP, respectively, and (ii) slight conformational changes of other active site residues, in particular Trp108, the base stacking residue with the pyridine cofactor moiety. This active site perturbation results in a mispositioning of the AGT-pyridoxamine 5'-phosphate (PMP) complex and of the external aldimine, as predicted by molecular modeling studies. Taken together, both predicted and observed movements caused by the S187F mutation are consistent with the following functional properties of the variant: (i) a 300- to 500-fold decrease in both the rate constant of L-alanine half-transamination and the kcat of the overall transamination, (ii) a different PMP binding mode and affinity, and (iii) a different microenvironment of the external aldimine. Proposals for the treatment of patients bearing S187F mutation are discussed on the basis of these results.

  3. Aspartate aminotransferase (AST) blood test

    MedlinePlus

    ... 2016:chap 73. Read More Acute kidney failure Acute pancreatitis Alanine transaminase (ALT) blood test ALP - blood test Burns Cardiac catheterization Enzyme Heart attack Hemolytic anemia Hepatic Liver cancer - hepatocellular carcinoma Liver ...

  4. From Genotype to Phenotype: Nonsense Variants in SLC13A1 Are Associated with Decreased Serum Sulfate and Increased Serum Aminotransferases

    PubMed Central

    Tise, Christina G.; Perry, James A.; Anforth, Leslie E.; Pavlovich, Mary A.; Backman, Joshua D.; Ryan, Kathleen A.; Lewis, Joshua P.; O’Connell, Jeffrey R.; Yerges-Armstrong, Laura M.; Shuldiner, Alan R.

    2016-01-01

    Using genomic applications to glean insights into human biology, we systematically searched for nonsense single nucleotide variants (SNVs) that are rare in the general population but enriched in the Old Order Amish (Amish) due to founder effect. We identified two nonlinked, nonsense SNVs (R12X and W48X) in SLC13A1 (allele frequencies 0.29% and 0.74% in the Amish; enriched 1.2-fold and 3.7-fold, compared to the outbred Caucasian population, respectively). SLC13A1 encodes the apical sodium-sulfate cotransporter (NaS1) responsible for sulfate (re)absorption in the kidneys and intestine. SLC13A1 R12X and W48X were independently associated with a 27.6% (P = 2.7 × 10−8) and 27.3% (P = 6.9 × 10−14) decrease in serum sulfate, respectively (P = 8.8 × 10-20 for carriers of either SLC13A1 nonsense SNV). We further performed the first exome- and genome-wide association study (ExWAS/GWAS) of serum sulfate and identified a missense variant (L348P) in SLC26A1, which encodes the basolateral sulfate-anion transporter (Sat1), that was associated with decreased serum sulfate (P = 4.4 × 10−12). Consistent with sulfate’s role in xenobiotic detoxification and protection against acetaminophen-induced hepatotoxicity, SLC13A1 nonsense SNV carriers had higher aminotransferase levels compared to noncarriers. Furthermore, SLC26A1 L348P was associated with lower whole-body bone mineral density (BMD) and higher serum calcium, consistent with the osteochondrodysplasia exhibited by dogs and sheep with naturally occurring, homozygous, loss-of-function mutations in Slc13a1. This study demonstrates the power and translational potential of systematic identification and characterization of rare, loss-of-function variants and warrants additional studies to better understand the importance of sulfate in human physiology, disease, and drug toxicity. PMID:27412988

  5. 3T3 fibroblasts transfected with a cDNA for mitochondrial aspartate aminotransferase express plasma membrane fatty acid-binding protein and saturable fatty acid uptake.

    PubMed Central

    Isola, L M; Zhou, S L; Kiang, C L; Stump, D D; Bradbury, M W; Berk, P D

    1995-01-01

    To explore the relationship between mitochondrial aspartate aminotransferase (mAspAT; EC 2.6.1.1) and plasma membrane fatty acid-binding protein (FABPpm) and their role in cellular fatty acid uptake, 3T3 fibroblasts were cotransfected with plasmid pMAAT2, containing a full-length mAspAT cDNA downstream of a Zn(2+)-inducible metallothionein promoter, and pFR400, which conveys methotrexate resistance. Transfectants were selected in methotrexate, cloned, and exposed to increasing methotrexate concentrations to induce gene amplification. Stably transfected clones were characterized by Southern blotting; those with highest copy numbers of pFR400 alone (pFR400) or pFR400 and pMAAT2 (pFR400/pMAAT2) were expanded for further study. [3H]Oleate uptake was measured in medium containing 500 microM bovine serum albumin and 125-1000 microM total oleate (unbound oleate, 18-420 nM) and consisted of saturable and nonsaturable components. pFR400/pMAAT2 cells exhibited no increase in the rate constant for nonsaturable oleate uptake or in the uptake rate of [14C]octanoate under any conditions. By contrast, Vmax (fmol/sec per 50,000 cells) of the saturable oleate uptake component increased 3.5-fold in pFR400/pMAAT2 cells compared to pFR400, with a further 3.2-fold increase in the presence of Zn2+. Zn2+ had no effect in pFR400 controls (P > 0.5). The overall increase in Vmax between pFR400 and pFR400/pMAAT2 in the presence of Zn2+ was 10.4-fold (P < 0.01) and was highly correlated (r = 0.99) with expression of FABPpm in plasma membranes as determined by Western blotting. Neither untransfected 3T3 nor pFR400 cells expressed cell surface FABPpm detectable by immunofluorescence. By contrast, plasma membrane immunofluorescence was detected in pFR400/pMAAT2 cells, especially if cultured in 100 microM Zn2+. The data support the dual hypotheses that mAspAT and FABPpm are identical and mediate saturable long-chain free fatty acid uptake. Images Fig. 1 Fig. 2 Fig. 4 Fig. 5 PMID:7568234

  6. Alanine-glyoxylate aminotransferase 2 (AGXT2) polymorphisms have considerable impact on methylarginine and β-aminoisobutyrate metabolism in healthy volunteers.

    PubMed

    Kittel, Anja; Müller, Fabian; König, Jörg; Mieth, Maren; Sticht, Heinrich; Zolk, Oliver; Kralj, Ana; Heinrich, Markus R; Fromm, Martin F; Maas, Renke

    2014-01-01

    Elevated plasma concentrations of asymmetric (ADMA) and symmetric (SDMA) dimethylarginine have repeatedly been linked to adverse clinical outcomes. Both methylarginines are substrates of alanine-glyoxylate aminotransferase 2 (AGXT2). It was the aim of the present study to simultaneously investigate the functional relevance and relative contributions of common AGXT2 single nucleotide polymorphisms (SNPs) to plasma and urinary concentrations of methylarginines as well as β-aminoisobutyrate (BAIB), a prototypic substrate of AGXT2. In a cohort of 400 healthy volunteers ADMA, SDMA and BAIB concentrations were determined in plasma and urine using HPLC-MS/MS and were related to the coding AGXT2 SNPs rs37369 (p.Val140Ile) and rs16899974 (p.Val498Leu). Volunteers heterozygous or homozygous for the AGXT2 SNP rs37369 had higher SDMA plasma concentrations by 5% and 20% (p = 0.002) as well as higher BAIB concentrations by 54% and 146%, respectively, in plasma and 237% and 1661%, respectively, in urine (both p<0.001). ADMA concentrations were not affected by both SNPs. A haplotype analysis revealed that the second investigated AGXT2 SNP rs16899974, which was not significantly linked to the other AGXT2 SNP, further aggravates the effect of rs37369 with respect to BAIB concentrations in plasma and urine. To investigate the impact of the amino acid exchange p.Val140Ile, we established human embryonic kidney cell lines stably overexpressing wild-type or mutant (p.Val140Ile) AGXT2 protein and assessed enzyme activity using BAIB and stable-isotope labeled [²H₆]-SDMA as substrate. In vitro, the amino acid exchange of the mutant protein resulted in a significantly lower enzyme activity compared to wild-type AGXT2 (p<0.05). In silico modeling of the SNPs indicated reduced enzyme stability and substrate binding. In conclusion, SNPs of AGXT2 affect plasma as well as urinary BAIB and SDMA concentrations linking methylarginine metabolism to the common genetic trait of hyper

  7. Alanine-glyoxylate aminotransferase 2 (AGXT2) Polymorphisms Have Considerable Impact on Methylarginine and β-aminoisobutyrate Metabolism in Healthy Volunteers

    PubMed Central

    König, Jörg; Mieth, Maren; Sticht, Heinrich; Zolk, Oliver; Kralj, Ana; Heinrich, Markus R.; Fromm, Martin F.; Maas, Renke

    2014-01-01

    Elevated plasma concentrations of asymmetric (ADMA) and symmetric (SDMA) dimethylarginine have repeatedly been linked to adverse clinical outcomes. Both methylarginines are substrates of alanine-glyoxylate aminotransferase 2 (AGXT2). It was the aim of the present study to simultaneously investigate the functional relevance and relative contributions of common AGXT2 single nucleotide polymorphisms (SNPs) to plasma and urinary concentrations of methylarginines as well as β-aminoisobutyrate (BAIB), a prototypic substrate of AGXT2. In a cohort of 400 healthy volunteers ADMA, SDMA and BAIB concentrations were determined in plasma and urine using HPLC-MS/MS and were related to the coding AGXT2 SNPs rs37369 (p.Val140Ile) and rs16899974 (p.Val498Leu). Volunteers heterozygous or homozygous for the AGXT2 SNP rs37369 had higher SDMA plasma concentrations by 5% and 20% (p = 0.002) as well as higher BAIB concentrations by 54% and 146%, respectively, in plasma and 237% and 1661%, respectively, in urine (both p<0.001). ADMA concentrations were not affected by both SNPs. A haplotype analysis revealed that the second investigated AGXT2 SNP rs16899974, which was not significantly linked to the other AGXT2 SNP, further aggravates the effect of rs37369 with respect to BAIB concentrations in plasma and urine. To investigate the impact of the amino acid exchange p.Val140Ile, we established human embryonic kidney cell lines stably overexpressing wild-type or mutant (p.Val140Ile) AGXT2 protein and assessed enzyme activity using BAIB and stable-isotope labeled [2H6]-SDMA as substrate. In vitro, the amino acid exchange of the mutant protein resulted in a significantly lower enzyme activity compared to wild-type AGXT2 (p<0.05). In silico modeling of the SNPs indicated reduced enzyme stability and substrate binding. In conclusion, SNPs of AGXT2 affect plasma as well as urinary BAIB and SDMA concentrations linking methylarginine metabolism to the common genetic trait of hyper

  8. Biochemical characterization of the Campylobacter jejuni Cj1294, a novel UDP-4-keto-6-deoxy-GlcNAc aminotransferase that generates UDP-4-amino-4,6-dideoxy-GalNAc.

    PubMed

    Obhi, Ravinder Kaur; Creuzenet, Carole

    2005-05-27

    Campylobacter jejuni produces multiple glycoproteins whose glycans contain 4-amino 6-deoxy sugars or their derivatives, such as diacetamidobacillosamine or pseudaminic acid. Because the proteoglycans contribute to bacterial virulence and their constitutive sugars are not commonly found in humans, inhibitors developed against the enzymes that are responsible for their biosynthesis could be novel therapeutic targets to fight this important food-borne pathogen. The biosynthesis of diacetamidobacillosamine is anticipated to involve a sugar nucleotide C6 dehydratase, a C4 aminotransferase and an acetyltransferase. We have identified a set of genes (cj1293, cj1294, and cj1298) potentially encoding one of each enzymatic activity, and demonstrated earlier that Cj1293 was a UDP-GlcNAc-specific C6 dehydratase. Others have shown that Cj1293 was involved in protein glycosylation. Here, we report on our investigation of the potential activity of Cj1294 as a sugar nucleotide C4 aminotransferase. Our biochemical characterization of overexpressed and purified protein shows that Cj1294 is a pyridoxal phosphate-dependent aminotransferase specific for UDP-4-keto-6-deoxy-GlcNAc that uses preferentially glutamic acid as an amino donor. A detailed physicokinetic study of Cj1294 was performed to determine the K(m) of 1.28 +/- 0.2 mm and k(cat) of 11.5 +/- 1.3 min(-1). Also, two residues essential for protein stability and activity, Arg(228) and Lys(181), respectively, were identified by site-directed mutagenesis. Finally, we demonstrated by NMR analysis of purified reaction product that Cj1294 produces UDP-4-amino-4,6-dideoxy-GalNAc. These results indicate that Cj1294 is involved in the biosynthesis of diacetamidofucosamine, a C4 epimer of diacetamidobacillosamine not yet described in C. jejuni proteoglycans, suggesting that the composition of C. jejuni proteoglycans is more variable than anticipated.

  9. Genomic and Biochemical Analysis of the Diaminopimelate and Lysine Biosynthesis Pathway in Verrucomicrobium spinosum: Identification and Partial Characterization of L,L-Diaminopimelate Aminotransferase and UDP-N-Acetylmuramoylalanyl-D-glutamyl-2,6-meso-Diaminopimelate Ligase.

    PubMed

    Nachar, Victoria R; Savka, Francisco C; McGroty, Sean E; Donovan, Katherine A; North, Rachel A; Dobson, Renwick C J; Buckley, Larry J; Hudson, André O

    2012-01-01

    The Gram-negative bacterium Verrucomicrobium spinosum has attracted interest in recent years following the sequencing and annotation of its genome. Comparative genomic analysis of V. spinosum using diaminopimelate/lysine metabolic genes from Chlamydia trachomatis suggests that V. spinosum employs the L,L-diaminopimelate aminotransferase (DapL) pathway for diaminopimelate/lysine biosynthesis. The open reading frame corresponding to the putative dapL ortholog was cloned and the recombinant enzyme was shown to possess L,L-diaminopimelate aminotransferase activity in vitro. In vivo analysis using functional complementation confirmed that the dapL ortholog was able to functionally complement an E. coli mutant that confers auxotrophy for diaminopimelate and lysine. In addition to its role in lysine biosynthesis, the intermediate diaminopimelate has an integral role in peptidoglycan biosynthesis. To this end, the UDP-N-acetylmuramoylalanyl-d-glutamyl-2,6-meso-diaminopimelate ligase ortholog was also identified, cloned, and was shown to possess meso-diaminopimelate ligase activity in vivo. The L,L-diaminopimelate aminotransferase pathway has been experimentally confirmed in several bacteria, some of which are deemed pathogenic to animals. Since animals, and particularly humans, lack the genetic machinery for the synthesis of diaminopimelate/lysine de novo, the enzymes involved in this pathway are attractive targets for development of antibiotics. Whether dapL is an essential gene in any bacteria is currently not known. V. spinosum is an excellent candidate to investigate the essentiality of dapL, since the bacterium employs the DapL pathway for lysine and cell wall biosynthesis, is non-pathogenic to humans, facile to grow, and can be genetically manipulated.

  10. Genomic and Biochemical Analysis of the Diaminopimelate and Lysine Biosynthesis Pathway in Verrucomicrobium spinosum: Identification and Partial Characterization of L,L-Diaminopimelate Aminotransferase and UDP-N-Acetylmuramoylalanyl-D-glutamyl-2,6-meso-Diaminopimelate Ligase

    PubMed Central

    Nachar, Victoria R.; Savka, Francisco C.; McGroty, Sean E.; Donovan, Katherine A.; North, Rachel A.; Dobson, Renwick C. J.; Buckley, Larry J.; Hudson, André O.

    2012-01-01

    The Gram-negative bacterium Verrucomicrobium spinosum has attracted interest in recent years following the sequencing and annotation of its genome. Comparative genomic analysis of V. spinosum using diaminopimelate/lysine metabolic genes from Chlamydia trachomatis suggests that V. spinosum employs the L,L-diaminopimelate aminotransferase (DapL) pathway for diaminopimelate/lysine biosynthesis. The open reading frame corresponding to the putative dapL ortholog was cloned and the recombinant enzyme was shown to possess L,L-diaminopimelate aminotransferase activity in vitro. In vivo analysis using functional complementation confirmed that the dapL ortholog was able to functionally complement an E. coli mutant that confers auxotrophy for diaminopimelate and lysine. In addition to its role in lysine biosynthesis, the intermediate diaminopimelate has an integral role in peptidoglycan biosynthesis. To this end, the UDP-N-acetylmuramoylalanyl-d-glutamyl-2,6-meso-diaminopimelate ligase ortholog was also identified, cloned, and was shown to possess meso-diaminopimelate ligase activity in vivo. The L,L-diaminopimelate aminotransferase pathway has been experimentally confirmed in several bacteria, some of which are deemed pathogenic to animals. Since animals, and particularly humans, lack the genetic machinery for the synthesis of diaminopimelate/lysine de novo, the enzymes involved in this pathway are attractive targets for development of antibiotics. Whether dapL is an essential gene in any bacteria is currently not known. V. spinosum is an excellent candidate to investigate the essentiality of dapL, since the bacterium employs the DapL pathway for lysine and cell wall biosynthesis, is non-pathogenic to humans, facile to grow, and can be genetically manipulated. PMID:22783236

  11. Expression, purification, crystallization, data collection and preliminary biochemical characterization of methicillin-resistant Staphylococcus aureus Sar2028, an aspartate/tyrosine/phenylalanine pyridoxal-5′-phosphate-dependent aminotransferase

    SciTech Connect

    Seetharamappa, Jaldappagari; Oke, Muse; Liu, Huanting; McMahon, Stephen A.; Johnson, Kenneth A.; Carter, Lester; Dorward, Mark; Zawadzki, Michal; Overton, Ian M.; Niekirk, C. A. Johannes van; Graham, Shirley; Botting, Catherine H.; Taylor, Garry L.; White, Malcolm F.; Barton, Geoffrey J.; Coote, Peter J.; Naismith, James H.

    2007-05-01

    As part of work on S. aureus, the crystallization of Sar2028, a protein that is upregulated in MRSA, is reported. Sar2028, an aspartate/tyrosine/phenylalanine pyridoxal-5′-phosphate-dependent aminotransferase with a molecular weight of 48 168 Da, was overexpressed in methicillin-resistant Staphylococcus aureus compared with a methicillin-sensitive strain. The protein was expressed in Escherichia coli, purified and crystallized. The protein crystallized in a primitive orthorhombic Laue group with unit-cell parameters a = 83.6, b = 91.3, c = 106.0 Å, α = β = γ = 90°. Analysis of the systematic absences along the three principal axes indicated the space group to be P2{sub 1}2{sub 1}2{sub 1}. A complete data set was collected to 2.5 Å resolution.

  12. Expression, purification, crystallization, data collection and preliminary biochemical characterization of methicillin-resistant Staphylococcus aureus Sar2028, an aspartate/tyrosine/phenylalanine pyridoxal-5′-phosphate-dependent aminotransferase

    PubMed Central

    Seetharamappa, Jaldappagari; Oke, Muse; Liu, Huanting; McMahon, Stephen A.; Johnson, Kenneth A.; Carter, Lester; Dorward, Mark; Zawadzki, Michal; Overton, Ian M.; van Niekirk, C. A. Johannes; Graham, Shirley; Botting, Catherine H.; Taylor, Garry L.; White, Malcolm F.; Barton, Geoffrey J.; Coote, Peter J.; Naismith, James H.

    2007-01-01

    Sar2028, an aspartate/tyrosine/phenylalanine pyridoxal-5′-phosphate-dependent aminotransferase with a molecular weight of 48 168 Da, was overexpressed in methicillin-resistant Staphylococcus aureus compared with a methicillin-sensitive strain. The protein was expressed in Escherichia coli, purified and crystallized. The protein crystallized in a primitive orthorhombic Laue group with unit-cell parameters a = 83.6, b = 91.3, c = 106.0 Å, α = β = γ = 90°. Analysis of the systematic absences along the three principal axes indicated the space group to be P212121. A complete data set was collected to 2.5 Å resolution. PMID:17565195

  13. High light inhibits chlorophyll biosynthesis at the level of 5-aminolevulinate synthesis during de-etiolation in cucumber (Cucumis sativus) cotyledons.

    PubMed

    Aarti, D; Tanaka, R; Ito, H; Tanaka, A

    2007-01-01

    Using the vascular plant Cucumis sativus (cucumber) as a model, we studied the effects of high (intense and excess) light upon chlorophyll biosynthesis during de-etiolation. When illuminated with high light (1500-1600 microE/m2/s), etiolated cucumber cotyledons failed to synthesize chlorophyll entirely. However, upon transfer to low light conditions (40-45 microE/m2/s), chlorophyll biosynthesis and subsequent accumulation resumed following an initial 2-12 h delay. Duration of high light treatment negatively correlated with chlorophyll biosynthetic activity. Specifically, we found that high light severely inhibited 5-aminolevulinic acid (ALA) synthesis. This effect partly could be because of the decrease in protein level of glutamyl-tRNA reductase (GluTR) observed. Protein level of glutamate-1-semialdehyde (GSA-AT) remained unchanged. It was also found that high light did not suppress HEMA 1 expression. Therefore, we speculated that this significant inhibition of ALA synthesis might have occurred mainly because of concomitant inactivation of GluTR and/or inhibition of complex formation between GluTR and GSA-AT. Our further observation that both methyl viologen and rose bengal similarly inhibit ALA synthesis under low light conditions suggested that reactive oxygen species (ROS) could be responsible for the inhibition of ALA synthesis in cotyledons exposed to high light conditions.

  14. Effects of test spills of chemically dispersed and nondispersed oil on the activity of aspartate aminotransferase and glucose-6-phosphate dehydrogenase in two intertidal bivalves, Mya arenaria and Mytilus edulis

    SciTech Connect

    Gilfillan, E.S.; Foster, J.; Gerber, R.; Hanson, S.A.; Page, D.S.; Vallas, D.

    1982-10-01

    In 1981, two test oil spills were made in Maine. One spill was 975 L (250 gal) of Murban crude oil; the other was 975 L of Murban crude oil premixed with 97 L (25 gal) of Corexit 9527. The uptake of the oil and its effects on enzymatic activity in two species of common intertidal bivalve mollusks, Mya arenaria and Mytilus edulis, were studied. Data were obtained on uptake and depuration of the oil for each species; data were also obtained on the activity of glucose-6-phosphate dehydrogenase and aspartate aminotransferase for each species. Data were collected both before and after each of the spills. Much less oil was taken up by the populations of animals exposed to chemically dispersed oil than by those exposed to nondispersed oil. Rates of depuration were the same for each species; they were also the same regardless of oil exposure. Significant long-term effects on enzyme activity were detected only in those animals exposed to nondispersed oil.

  15. Rice Stress Associated Protein 1 (OsSAP1) Interacts with Aminotransferase (OsAMTR1) and Pathogenesis-Related 1a Protein (OsSCP) and Regulates Abiotic Stress Responses

    PubMed Central

    Kothari, Kamakshi S.; Dansana, Prasant K.; Giri, Jitender; Tyagi, Akhilesh K.

    2016-01-01

    Stress associated proteins (SAPs) are the A20/AN1 zinc-finger containing proteins which can regulate the stress signaling in plants. The rice SAP protein, OsSAP1 has been shown to confer abiotic stress tolerance to plants, when overexpressed, by modulating the expression of endogenous stress-related genes. To further understand the mechanism of OsSAP1-mediated stress signaling, OsSAP1 interacting proteins were identified using yeast two-hybrid analysis. Two novel proteins, aminotransferase (OsAMTR1) and a SCP/TAPS or pathogenesis-related 1 class of protein (OsSCP) were found to interact with OsSAP1. The genes encoding OsAMTR1 and OsSCP were stress-responsive and showed higher expression upon abiotic stress treatments. The role of OsAMTR1 and OsSCP under stress was analyzed by overexpressing them constitutively in Arabidopsis and responses of transgenic plants were assessed under salt and water-deficit stress. The OsAMTR1 and OsSCP overexpressing plants showed higher seed germination, root growth and fresh weight than wild-type plants under stress conditions. Overexpression of OsAMTR1 and OsSCP affected the expression of many known stress-responsive genes which were not affected by the overexpression of OsSAP1. Moreover, the transcript levels of OsSCP and OsAMTR1 were also unaffected by the overexpression of OsSAP1. Hence, it was concluded that OsSAP1 regulates the stress responsive signaling by interacting with these proteins which further regulate the downstream stress responsive gene expression. PMID:27486471

  16. Targeted deletion of the kynurenine aminotransferase ii gene reveals a critical role of endogenous kynurenic acid in the regulation of synaptic transmission via alpha7 nicotinic receptors in the hippocampus.

    PubMed

    Alkondon, Manickavasagon; Pereira, Edna F R; Yu, Ping; Arruda, Emerson Z; Almeida, Luis E F; Guidetti, Paolo; Fawcett, William P; Sapko, Michael T; Randall, William R; Schwarcz, Robert; Tagle, Danilo A; Albuquerque, Edson X

    2004-05-12

    It has been postulated that endogenous kynurenic acid (KYNA) modulates alpha7* nicotinic acetylcholine receptor (nAChR) and NMDA receptor activities in the brain.a To test this hypothesis, alpha7* nAChR and NMDA receptor functions were studied in mice with a targeted null mutation in the gene encoding kynurenine aminotransferase II (mKat-2-/- mice), an enzyme responsible for brain KYNA synthesis. At 21 postnatal days, mKat-2-/- mice had lower hippocampal KYNA levels and higher spontaneous locomotor activity than wild-type (WT) mice. At this age, alpha7* nAChR activity induced by exogenous application of agonists to CA1 stratum radiatum interneurons was approximately 65% higher in mKat-2-/- than WT mice. Binding studies indicated that the enhanced receptor activity may not have resulted from an increase in alpha7* nAChR number. In 21-d-old mKat-2-/- mice, endogenous alpha7* nAChR activity in the hippocampus was also increased, leading to an enhancement of GABAergic activity impinging onto CA1 pyramidal neurons that could be reduced significantly by acute exposure to KYNA (100 nM). The activities of GABA(A) and NMDA receptors in the interneurons and of alpha3beta4* nAChRs regulating glutamate release onto these neurons were comparable between mKat-2-/- and WT mice. By 60 d of age, KYNA levels and GABAergic transmission in the hippocampus and locomotor activity were similar between mKat-2-/- and WT mice. Our findings that alpha7* nAChRs are major targets for KYNA in the brain may provide insights into the pathophysiology of schizophrenia and Alzheimer's disease, disorders in which brain KYNA levels are increased and alpha7* nAChR functions are impaired.

  17. Mechanism of Inactivation of γ-Aminobutyric Acid Aminotransferase by (1S ,3S)-3-Amino-4-difluoromethylene-1-cyclopentanoic Acid (CPP-115)

    SciTech Connect

    Lee, Hyunbeom; Doud, Emma H.; Wu, Rui; Sanishvili, Ruslan; Juncosa, Jose I.; Liu, Dali; Kelleher, Neil L.; Silverman, Richard B.

    2015-01-23

    γ-Aminobutyric acid aminotransferase (GABA-AT) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that degrades GABA, the principal inhibitory neurotransmitter in mammalian cells. When the concentration of GABA falls below a threshold level, convulsions can occur. Inhibition of GABA-AT raises GABA levels in the brain, which can terminate seizures as well as have potential therapeutic applications in treating other neurological disorders, including drug addiction. Among the analogues that we previously developed, (1S,3S)-3-amino-4-difluoromethylene-1-cyclopentanoic acid (CPP-115) showed 187 times greater potency than that of vigabatrin, a known inactivator of GABA-AT and approved drug (Sabril) for the treatment of infantile spasms and refractory adult epilepsy. Recently, CPP-115 was shown to have no adverse effects in a Phase I clinical trial. Here we report a novel inactivation mechanism for CPP-115, a mechanism-based inactivator that undergoes GABA-AT-catalyzed hydrolysis of the difluoromethylene group to a carboxylic acid with concomitant loss of two fluoride ions and coenzyme conversion to pyridoxamine 5'-phosphate (PMP). The partition ratio for CPP-115 with GABA-AT is about 2000, releasing cyclopentanone-2,4-dicarboxylate (22) and two other precursors of this compound (20 and 21). Time-dependent inactivation occurs by a conformational change induced by the formation of the aldimine of 4-aminocyclopentane-1,3-dicarboxylic acid and PMP (20), which disrupts an electrostatic interaction between Glu270 and Arg445 to form an electrostatic interaction between Arg445 and the newly formed carboxylate produced by hydrolysis of the difluoromethylene group in CPP-115, resulting in a noncovalent, tightly bound complex. Ultimately, this represents a novel mechanism for inactivation of GABA-AT and a new approach for the design of mechanism-based inactivators in general.

  18. Liver biomarker and in vitro assessment confirm the hepatic origin of aminotransferase elevations lacking histopathological correlate in beagle dogs treated with GABA{sub A} receptor antagonist NP260

    SciTech Connect

    Harrill, Alison H.; Eaddy, John S.; Rose, Kelly; Cullen, John M.; Ramanathan, Lakshmi; Wanaski, Stephen; Collins, Stephen; Ho, Yu; Watkins, Paul B.; LeCluyse, Edward L.

    2014-06-01

    NP260 was designed as a first-in-class selective antagonist of α4-subtype GABA{sub A} receptors that had promising efficacy in animal models of pain, epilepsy, psychosis, and anxiety. However, development of NP260 was complicated following a 28-day safety study in dogs in which pronounced elevations of serum aminotransferase levels were observed, although there was no accompanying histopathological indication of hepatocellular injury. To further investigate the liver effects of NP260, we assayed stored serum samples from the 28-day dog study for liver specific miRNA (miR-122) as well as enzymatic biomarkers glutamate dehydrogenase and sorbitol dehydrogenase, which indicate liver necrosis. Cytotoxicity assessments were conducted in hepatocytes derived from dog, rat, and human liver samples to address the species specificity of the liver response to NP260. All biomarkers, except ALT, returned toward baseline by Day 29 despite continued drug treatment, suggesting adaptation to the initial injury. In vitro analysis of the toxicity potential of NP260 to primary hepatocytes indicated a relative sensitivity of dog > human > rat, which may explain, in part, why the liver effects were not evident in the rodent safety studies. Taken together, the data indicate that a diagnostic biomarker approach, coupled with sensitive in vitro screening strategies, may facilitate interpretation of toxicity potential when an adaptive event masks the underlying toxicity. - Highlights: • NP260 caused ALT elevations in dogs without evidence of hepatocellular injury. • SDH, GLDH, and miRNA-122 elevations occurred, confirming hepatocellular necrosis. • NP260 toxicity is greater in dog and human hepatocytes than in rat hepatocytes. • Species sensitivity may explain why the rodent studies failed to indicate risk. • Diagnostic biomarkers and hepatocyte studies aid interpretation of hepatotoxicity.

  19. Evolution of alanine:glyoxylate aminotransferase 1 peroxisomal and mitochondrial targeting. A survey of its subcellular distribution in the livers of various representatives of the classes Mammalia, Aves and Amphibia.

    PubMed

    Danpure, C J; Fryer, P; Jennings, P R; Allsop, J; Griffiths, S; Cunningham, A

    1994-08-01

    As part of a wider study on the molecular evolution of alanine:glyoxylate aminotransferase 1 (AGT1) intracellular compartmentalization, we have determined the subcellular distribution of immunoreactive AGT1, using postembedding protein A-gold immunoelectron microscopy, in the livers of various members of the classes Mammalia, Aves, and Amphibia. As far as organellar distribution is concerned, three categories could be distinguished. In members of the first category (type I), all, or nearly all, of the immunoreactive AGT1 was concentrated within the peroxisomes. In the second category (type II), AGT1 was found more evenly distributed in both peroxisomes and mitochondria. In the third category (type III), AGT1 was localized mainly within the mitochondria with much lower, but widely variable, amounts in the peroxisomes. Type I animals include the human, two great apes (gorilla, orangutan), two Old World monkeys (anubis baboon, Japanese macaque), a New World monkey (white-faced Saki monkey), a lago, morph (European rabbit), a bat (Seba's short-tailed fruit bat), two caviomorph rodents (guinea pig, orange-rumped agouti), and two Australian marsupials (koala, Bennett's wallaby). Type II animals include two New World monkeys (common marmoset, cotton-top tamarin), three prosimians (brown lemur, fat-tailed dwarf lemur, pygmy slow loris), five rodents (a hybrid crested porcupine, Colombian ground squirrel, laboratory rat, laboratory mouse, golden hamster), an American marsupial (grey short-tailed opossum), and a bird (raven). Type III animals include the large tree shrew, three insectivores (common Eurasian mole, European hedgehog, house shrew), four carnivores (domestic cat, ocelot, domestic dog, polecat ferret), and an amphibian (common frog). In addition to these categories, some animals (e.g. guinea pig, common frog) possessed significant amounts of cytosolic AGT1. Whereas the subcellular distribution of AGT1 in some orders (e.g. Insectivora and Carnivora) did not appear

  20. Antiretroviral Drugs and Risk of Chronic Alanine Aminotransferase Elevation in Human Immunodeficiency Virus (HIV)-Monoinfected Persons: The Data Collection on Adverse Events of Anti-HIV Drugs Study

    PubMed Central

    Kovari, Helen; Sabin, Caroline A.; Ledergerber, Bruno; Ryom, Lene; Reiss, Peter; Law, Matthew; Pradier, Christian; Dabis, Francois; d'Arminio Monforte, Antonella; Smith, Colette; de Wit, Stephane; Kirk, Ole; Lundgren, Jens D.; Weber, Rainer

    2016-01-01

    Background. Although human immunodeficiency virus (HIV)-positive persons on antiretroviral therapy (ART) frequently have chronic liver enzyme elevation (cLEE), the underlying cause is often unclear. Methods. Data Collection on Adverse Events of Anti-HIV Drugs (D:A:D) Study participants without chronic viral hepatitis were observed to the earliest of cLEE (elevated aminotransferase ≥6 months), death, last follow-up, or January 2, 2014. Antiretroviral treatment exposure was categorized as follows: no exposure and ongoing short- and long-term exposure (<2 or ≥2 years) after initiation. Association between development of cLEE and ART exposure was investigated using Poisson regression. Results. Among 21 485 participants observed for 105 413 person-years (PY), 6368 developed cLEE (incidence 6.04/100 PY; 95% confidence interval [CI], 5.89–6.19). Chronic liver enzyme elevation was associated with short-and long-term exposure to didanosine (<2 years rate ratio [RR] = 1.29, 95% CI, 1.11–1.49; >2 years RR = 1.26, 95% CI, 1.13–1.41); stavudine (<2 years RR = 1.51, 95% CI, 1.26–1.81; >2 years RR = 1.17, 95% CI, 1.03–1.32), and tenofovir disoproxil fumarate (<2 years RR = 1.55, 95% CI, 1.40–1.72; >2 years RR = 1.18, 95% CI, 1.05–1.32), but only short-term exposure to nevirapine (<2 years RR = 1.44, 95% CI, 1.29–1.61), efavirenz (<2 years RR = 1.14, 95% CI, 1.03–1.26), emtricitabine (<2 years RR = 1.18, 95% CI, 1.04–1.33), and atazanavir (<2 years RR = 1.20, 95% CI, 1.04–1.38). Chronic liver enzyme elevation was not associated with use of lamivudine, abacavir, and other protease inhibitors. Mortality did not differ between participants with and without cLEE. Conclusions. Although didanosine, stavudine, nevirapine, and efavirenz have been described to be hepatotoxic, we additionally observed a consistent association between tenofovir and cLEE emerging within the first 2 years after drug initiation. This novel tenofovir-cLEE signal should be

  1. Formation of the chlorophyll precursor. gamma. -aminolevulinic acid in cyanobacteria requires aminoacylation of a tRNA sup Glu species. [Synechocystis

    SciTech Connect

    O'Nell, G.P.; Peterson, D.M.; Schoen, A., Chen, Minwei; Soell, D. )

    1988-09-01

    In the chloroplasts of higher plants and algae, the biosynthesis of the chlorophyll precursor {gamma}-aminolevulinic acid (ALA) involves at least three enzymes and a tRNA species. Here we demonstrate that in cell extracts of the unicellular cyanobacterium Synechocystis sp. strain PCC 6803 ALA was formed from glutamate in a series of reactions in which activation of glutamate by glutamyl-tRNA{sup Glu} formation was the first step. The activated glutamate was reduced by a dehydrogenase which displayed tRNA sequence specificity. Fractionation of strain 6803 tRNA by reverse-phase chromatography and polyacrylamide gel electrophoresis yielded two pure tRNA{sup Glu} species which stimulated ALA synthesis in vitro. These tRNAs had identical primary sequence but differed in the nucleotide modification of their anticodon. The 6803 tRNA{sup Glu} was similar to the sequence of tRNA{sup Glu} species or tRNA genes from Escherichia coli and from chloroplasts of Euglena gracilis and higher plants. Southern blot analysis revealed at least two tRNA{sup Glu} gene copies in the 6803 chromosome. A glutamate-1-semialdehyde aminotransferase, the terminal enzyme in the conversion of glutamate to ALA in chloroplasts, was detected in 6803 cell extracts by the conversion of glutamate-1-semialdehyde to ALA and by the inhibition of this reaction by gabaculin.

  2. The Effect of Artichoke Leaf Extract on Alanine Aminotransferase and Aspartate Aminotransferase in the Patients with Nonalcoholic Steatohepatitis

    PubMed Central

    Rangboo, Vajiheh; Noroozi, Mostafa; Zavoshy, Roza; Rezadoost, Seyed Amirmansoor; Mohammadpoorasl, Asghar

    2016-01-01

    Background. Based on recent basic and clinical investigations, the extract of artichoke (Cynara scolymus) leaf has been revealed to be used for hepatoprotective and cholesterol reducing purposes. We aimed to assess the therapeutic effects of artichoke on biochemical and liver biomarkers in patients with nonalcoholic steatohepatitis (NASH). Methods. In a randomized double blind clinical trial, 60 consecutive patients suffering NASH were randomly assigned to receive Cynara scolymus extract (as 6 tablets per day consisting of 2700 mg extract of the herb) as the intervention group or placebo as the control group for two months. Results. Comparing changes in study markers following interventions showed improvement in liver enzymes. The levels of triglycerides and cholesterol were significantly reduced in the group treated with Cynara scolymus when compared to placebo group. To compare the role of Cynara scolymus use with placebo in changes in study parameters, multivariate linear regression models were employed indicating higher improvement in liver enzymes and also lipid profile particularly triglycerides and total cholesterol following administration of Cynara scolymus in comparison with placebo use. Conclusion. This study sheds light on the potential hepatoprotective activity and hypolipidemic effect of Cynara scolymus in management of NASH. This clinical trial is registered in the IRCT, Iranian Registry of Clinical Trials, by number IRCT2014070218321N1. PMID:27293900

  3. The Effect of Artichoke Leaf Extract on Alanine Aminotransferase and Aspartate Aminotransferase in the Patients with Nonalcoholic Steatohepatitis.

    PubMed

    Rangboo, Vajiheh; Noroozi, Mostafa; Zavoshy, Roza; Rezadoost, Seyed Amirmansoor; Mohammadpoorasl, Asghar

    2016-01-01

    Background. Based on recent basic and clinical investigations, the extract of artichoke (Cynara scolymus) leaf has been revealed to be used for hepatoprotective and cholesterol reducing purposes. We aimed to assess the therapeutic effects of artichoke on biochemical and liver biomarkers in patients with nonalcoholic steatohepatitis (NASH). Methods. In a randomized double blind clinical trial, 60 consecutive patients suffering NASH were randomly assigned to receive Cynara scolymus extract (as 6 tablets per day consisting of 2700 mg extract of the herb) as the intervention group or placebo as the control group for two months. Results. Comparing changes in study markers following interventions showed improvement in liver enzymes. The levels of triglycerides and cholesterol were significantly reduced in the group treated with Cynara scolymus when compared to placebo group. To compare the role of Cynara scolymus use with placebo in changes in study parameters, multivariate linear regression models were employed indicating higher improvement in liver enzymes and also lipid profile particularly triglycerides and total cholesterol following administration of Cynara scolymus in comparison with placebo use. Conclusion. This study sheds light on the potential hepatoprotective activity and hypolipidemic effect of Cynara scolymus in management of NASH. This clinical trial is registered in the IRCT, Iranian Registry of Clinical Trials, by number IRCT2014070218321N1.

  4. Elevation of Alanine Aminotransferase Activity Occurs after Activation of the Cell-Death Signaling Initiated by Pattern-Recognition Receptors ‎but before Activation of Cytolytic Effectors in NK or CD8+ T Cells in the Liver During Acute HCV Infection

    PubMed Central

    Choi, Youkyung H.; Jin, Nancy; Kelly, Fiona; Sakthivel, SenthilKumar K.; Yu, Tianwei

    2016-01-01

    Pattern-recognition receptors (PRRs) promote host defenses against HCV infection by binding to their corresponding adapter molecules leading to the initiation of innate immune responses including cell death. We investigated the expression of PRR genes, biomarkers of liver cell-death, and T cell and NK cell activation/inhibition-related genes in liver and serum obtained from three experimentally infected chimpanzees with acute HCV infection, and analyzed the correlation between gene expression levels and clinical profiles. Our results showed that expression of hepatic RIG-I, TLR3, TLR7, 2OAS1, and CXCL10 mRNAs was upregulated as early as 7 days post-inoculation and peaked 12 to 83 days post-inoculation. All of the three HCV infected chimpanzees exhibited significant elevations of serum alanine aminotransferase (ALT) activity between 70 and 95 days after inoculation. Elevated levels of serum cytokeratin 18 (CK-18) and caspases 3 and 7 activity coincided closely with the rise of ALT activity, and were preceded by significant increases in levels of caspase 3 and caspase 7 mRNAs in the liver. Particularly we found that significant positive auto-correlations were observed between RIG-I, TLR3, CXCL10, 2OAS1, and PD-L1 mRNA and ALT activity at 3 to 12 days before the peak of ALT activity. However, we observed substantial negative auto-correlations between T cell and NK cell activation/inhibition-related genes and ALT activity at 5 to 32 days after the peak of ALT activity. Our results indicated cell death signaling is preceded by early induction of RIG-I, TLR3, 2OAS1, and CXCL10 mRNAs which leads to elevation of ALT activity and this signaling pathway occurs before the activation of NK and T cells during acute HCV infection. Our study suggests that PRRs and type I IFN response may play a critical role in development of liver cell injury related to viral clearance during acute HCV infection. PMID:27788241

  5. Tomato fruit photosynthesis is seemingly unimportant in primary metabolism and ripening but plays a considerable role in seed development.

    PubMed

    Lytovchenko, Anna; Eickmeier, Ira; Pons, Clara; Osorio, Sonia; Szecowka, Marek; Lehmberg, Kerstin; Arrivault, Stephanie; Tohge, Takayuki; Pineda, Benito; Anton, Maria Teresa; Hedtke, Boris; Lu, Yinghong; Fisahn, Joachim; Bock, Ralph; Stitt, Mark; Grimm, Bernhard; Granell, Antonio; Fernie, Alisdair R

    2011-12-01

    Fruit of tomato (Solanum lycopersicum), like those from many species, have been characterized to undergo a shift from partially photosynthetic to truly heterotrophic metabolism. While there is plentiful evidence for functional photosynthesis in young tomato fruit, the rates of carbon assimilation rarely exceed those of carbon dioxide release, raising the question of its role in this tissue. Here, we describe the generation and characterization of lines exhibiting a fruit-specific reduction in the expression of glutamate 1-semialdehyde aminotransferase (GSA). Despite the fact that these plants contained less GSA protein and lowered chlorophyll levels and photosynthetic activity, they were characterized by few other differences. Indeed, they displayed almost no differences in fruit size, weight, or ripening capacity and furthermore displayed few alterations in other primary or intermediary metabolites. Although GSA antisense lines were characterized by significant alterations in the expression of genes associated with photosynthesis, as well as with cell wall and amino acid metabolism, these changes were not manifested at the phenotypic level. One striking feature of the antisense plants was their seed phenotype: the transformants displayed a reduced seed set and altered morphology and metabolism at early stages of fruit development, although these differences did not affect the final seed number or fecundity. Taken together, these results suggest that fruit photosynthesis is, at least under ambient conditions, not necessary for fruit energy metabolism or development but is essential for properly timed seed development and therefore may confer an advantage under conditions of stress.

  6. Transcriptomic analysis for elucidating the physiological effects of 5-aminolevulinic acid accumulation on Corynebacterium glutamicum.

    PubMed

    Yu, Xiaoli; Jin, Haiying; Cheng, Xuelian; Wang, Qian; Qi, Qingsheng

    2016-11-01

    5-Aminolevulinic acid (ALA), the committed intermediate of the heme biosynthetic pathway, attracts close attention among researchers because of its potential applications to cancer treatment and agriculture. Overexpression of heterologous hemA and hemL, which encode glutamyl-tRNA reductase and glutamate-1-semialdehyde aminotransferase, respectively, in Corynebacterium glutamicum produces ALA, although whether ALA accumulation causes unintended effects on the host is unknown. Here we used an integrated systems approach to compare global transcriptional changes induced by the expression of hemA and hemL. Metabolic pathway such as glycolysis was inhibited, but tricarboxylic acid cycle, pentose phosphate pathway, and respiratory metabolism were stimulated. Moreover, the transcriptional levels of certain genes involved in heme biosynthesis were up-regulated, and the data implicate the two-component system (TCS) HrrSA was involved in the regulation of heme synthesis. With these understandings, it is proposed that ALA accumulation stimulates heme synthesis pathway and respiratory metabolism. Our study illuminates the physiological effects of overexpressing hemA and hemL on the phenotype of C. glutamicum and contributes important insights into the regulatory mechanisms of the heme biosynthetic pathways.

  7. Isolation, characterization, and expression analyses of tryptophan aminotransferase genes in a maize dek18 mutant

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The dek18 mutant of maize has decreased auxin content in kernels. Molecular and functional characterization of this mutant line offers the possibility to better understand auxin biology in maize seed development. Seeds of the dek18 mutants are smaller compared to wild type seeds and the vegetative d...

  8. An easy method for diagnosing macro-aspartate aminotransferase: a case series.

    PubMed

    Beşer, Omer Faruk; Laçinel, Sibel; Gülcü, Didem; Kutlu, Tufan; Cullu Çokuğraş, Fügen; Erkan, Tülay

    2014-10-01

    Macro-aspartate transaminase (macro-AST) must be considered when the aspartate transaminase (AST) level is chronically high without any liver, cardiac, or muscle disease. Many specialized laboratory techniques have been recommended for diagnosing macro-AST, including the polyethylene glycol immune precipitate technique, which is simple. This study presents a considerably easier method based on the studies of Davidson and Watson and Castiella et al. Our method is based on the decrease in the plasma AST level after storage of the macroenzyme at 2-8 °C for 5 days, and has the advantages of low cost, reliability, and practicality at any health center. In our eight cases of macro-AST, the AST activity at day 6 had decreased by more than 50% from day 1. This method is practical for primary healthcare facilities because of its easy application and accurate results, and obviated the need for unnecessary tests after diagnosis.

  9. Evidence for a Contribution of ALA Synthesis to Plastid-To-Nucleus Signaling

    SciTech Connect

    Czarnecki, Olaf; Gläßer, Christine; Chen, Jin-Gui; Mayer, Klaus F. X.; Grimm, Bernhard

    2012-01-01

    The formation of 5-aminolevulinic acid (ALA) in tetrapyrrole biosynthesis is widely controlled by environmental and metabolic feedback cues that determine the influx into the entire metabolic path. Because of its central role as the rate-limiting step, we hypothesized a potential role of ALA biosynthesis in tetrapyrrole-mediated retrograde signaling and exploited the direct impact of ALA biosynthesis on nuclear gene expression (NGE) by using two different approaches. Firstly, the Arabidopsis gun1, hy1 (gun2), hy2 (gun3), gun4 mutants showing uncoupled NGE from the physiological state of chloroplasts were thoroughly examined for regulatory modifications of ALA synthesis and transcriptional control in the nucleus. We found that reduced ALA-synthesizing capacity is common to analyzed gun mutants. Inhibition of ALA synthesis by gabaculine (GAB) that inactivates glutamate-1-semialdehyde aminotransferase and ALA feeding of wild-type and mutant seedlings corroborate the expression data of gun mutants. Transcript level of photosynthetic marker genes were enhanced in norflurazon (NF)-treated seedlings upon additional GAB treatment, while enhanced ALA amounts diminish these RNA levels in NF-treated wild-type in comparison to the solely NF-treated seedlings. Secondly, the impact of posttranslationally down-regulated ALA synthesis on NGE was investigated by global transcriptome analysis of GAB-treated Arabidopsis seedlings and the gun4-1 mutant, which is also characterized by reduced ALA formation. A common set of significantly modulated genes was identified indicating ALA synthesis as a potential signal emitter. The over-represented gene ontology categories of genes with decreased or increased transcript abundance highlight a few biological processes and cellular functions, which are remarkably affected in response to plastid-localized ALA biosynthesis. These results support the hypothesis that ALA biosynthesis correlates with retrograde signaling-mediated control of NGE.

  10. Decreased production of higher alcohols by Saccharomyces cerevisiae for Chinese rice wine fermentation by deletion of Bat aminotransferases.

    PubMed

    Zhang, Cui-Ying; Qi, Ya-Nan; Ma, Hong-Xia; Li, Wei; Dai, Long-Hai; Xiao, Dong-Guang

    2015-04-01

    An appropriate level of higher alcohols produced by yeast during the fermentation is one of the most important factors influencing Chinese rice wine quality. In this study, BAT1 and BAT2 single- and double-gene-deletion mutant strains were constructed from an industrial yeast strain RY1 to decrease higher alcohols during Chinese rice wine fermentation. The results showed that the BAT2 single-gene-deletion mutant strain produced best improvement in the production of higher alcohols while remaining showed normal growth and fermentation characteristics. Furthermore, a BAT2 single-gene-deletion diploid engineered strain RY1-Δbat2 was constructed and produced low levels of isobutanol and isoamylol (isoamyl alcohol and active amyl alcohol) in simulated fermentation of Chinese rice wine, 92.40 and 303.31 mg/L, respectively, which were 33.00 and 14.20 % lower than those of the parental strain RY1. The differences in fermentation performance between RY1-Δbat2 and RY1 were minor. Therefore, construction of this yeast strain is important in future development in Chinese wine industry and provides insights on generating yeast strains for other fermented alcoholic beverages.

  11. Determination of Alanine Aminotransferase with an Electrochemical Nano Ir-C Biosensor for the Screening of Liver Diseases

    PubMed Central

    Hsueh, Chang-Jung; Wang, Joanne H.; Dai, Liming; Liu, Chung-Chiun

    2011-01-01

    Alanine aminotransaminase (ALT), is an enzyme that normally resides in serum and body tissues, especially in the liver. It is released into the serum as a result of tissue injury; hence the concentration of ALT in the serum may be increased with acute damage to hepatic cells. A single use, disposable biosensor, comprising iridium nano-particle as catalyst dispersed on carbon paste, has been developed for the determination of ALT concentration. The biosensor is based on quantifying H2O2 concentration produced by a serial of ALT enzymatic reactions. It operates well at room temperature in different physiological fluids: phosphate buffer, calf serum and human serum for ALT concentration of 0–544 ng/mL. Experimental results in human serum are compared to those obtained by spectrophotometric assays with excellent agreement. Therefore, the Ir/C biosensor shows good relationship on the dilution of concentrated ALT clinical applications. PMID:25586923

  12. Development of an Electrochemical-Based Aspartate Aminotransferase Nanoparticle Ir-C Biosensor for Screening of Liver Diseases

    PubMed Central

    Hsueh, Chang-Jung (Alan); Wang, Joanne H.; Dai, Liming; Liu, Chung-Chiun

    2012-01-01

    Aspartate aminotransaminase (AST) is a hepatocelluar enzyme released into the bloodstream when hepatic cells are damaged, resulting in elevated blood levels of AST. A single use, disposable biosensor prototype, composed of catalytic iridium nano-particles dispersed on carbon paste, was developed to detect enzymatically-produced H2O2 in AST-mediated reactions. This biosensor is capable of measuring AST levels in a phosphate buffer and undiluted human serum over the concentration range of 0 to 0.89 μg/mL AST concentration (corresponding to 0–250 UL−1 specific activity). The biosensor operates at relatively low oxidation potential (+0.3 volt (V) versus the printed Ag/AgCl), minimizing any potential chemical interference in human serum. The measurements of AST in human serum using the biosensor compared well with those measured by standard hospital spectrophotometric assays. This Ir-C biosensor may be useful for AST measurements in the clinical environment. PMID:25585713

  13. Cannabinoid Receptor 2-63 QQ Variant Is Associated with Persistently Normal Aminotransferase Serum Levels in Chronic Hepatitis C

    PubMed Central

    Coppola, Nicola; Zampino, Rosa; Sagnelli, Caterina; Bellini, Giulia; Marrone, Aldo; Stanzione, Maria; Capoluongo, Nicolina; Boemio, Adriana; Minichini, Carmine; Adinolfi, Luigi Elio; Maione, Sabatino; Giudice, Emanuele Miraglia Del; Sagnelli, Evangelista; Rossi, Francesca

    2014-01-01

    Background and Aim To evaluate in anti-HCV-positive patients the clinical impact of the rs35761398 variant of the CNR2 gene leading to the substitution of Gln (Q) of codon 63 of the cannabinoid receptor 2 (CB2) with Arg (R). Patients and Methods 253 consecutive anti-HCV-/HCV-RNA-positive patients were enrolled, of whom 53 were HCV carriers with persistently normal ALT (PNALT group) and 200 had a history of steadily abnormal serum ALT values (abnormal ALT group). All patients were naive for antiviral therapy and were screened for the CNR2 rs35761398 polymorphism by a TaqMan assay. Results Subjects in the PNALT group, compared with those in the abnormal ALT group were older (58.5±12 vs. 50.7±12.4 years, p = 0.001), more frequently female (66% vs. 42%, p = 0.003), with lower body massindex (BMI) (24.5±3.1 vs. 26.6±4.6, p = 0.003), and more frequently with HCV genotype 2 (43.1% vs 17.7%, p = 0.0002) and CB2-63 QQ variant (34% vs. 11%, p = 0.0001). Considering all 253 patients, no difference in the demographic, biochemical, or virological data was observed between patients in the different CB2-63 variants. The logistic regression analysis identified CB2-63 QQ, HCV genotype 2, older age and lower BMI as independent predictors of PNALT (p<0.00001). Discussion The CB2-63 QQ variant in HCV patients was independently associated with the PNALT status. PMID:24940753

  14. [A young boy with elevated aminotransferases in physical examination--Two novel missense mutations associated with Wilson's disease were found].

    PubMed

    Zhu, Yu; Deng, Si-Yan; Wan, Chao-Min

    2015-07-01

    A 3-year-old boy had abnormal liver function, which was found in physical examination, for 5 months before admission. He had no symptoms such as anorexia, poor appetite, and jaundice, had normal growth and development, and showed no hepatosplenomegaly. Laboratory examination revealed significantly reduced ceruloplasmin (35 mg/L), as well as negative hepatotropic virus, cytomegalovirus, and Epstein-Barr virus. There were normal muscle enzymes, blood glucose, and blood ammonia and negative liver-specific autoantibodies. The boy had negative K-F ring and normal 24-hour urine copper (0.56 μmol/L). The ATP7B gene testing for the boy, his sister, and their parents detected two novel missense mutations in the boy and his sister, i.e., compound heterozygous mutations in exon 7 (c.2075T>C, p.L692P) and exon 13 (c.3044T>C, p.L1015P), which were inherited from their father and mother, respectively. Wilson's disease was confirmed by genetic diagnosis in the boy and his sister. The boy and his sister were given a low-copper diet. The boy was administered with penicillamine for decoppering and zinc supplement against copper uptake. His sister received zinc supplement alone because no clinical symptoms were observed. The boy showed normal liver function in the reexamination after 3 months of treatment.

  15. Coffee consumption is associated with lower serum aminotransferases in the general Korean population and in those at high risk for hepatic disease.

    PubMed

    Oh, Myueng Guen; Han, Mi Ah; Kim, Man Woo; Park, Chan Guk; Kim, Young Dae; Lee, Jun

    2016-12-01

    背景和目的:咖啡对肝酶的有利影响已在世界范围内被报道。该研究调查了 在韩国成年人的咖啡消费量和血清转氨酶浓度之间的关系。方法与研究设 计:数据来自第四和第五次韩国国家健康和营养检测普查。男性丙氨酸氨基 转移酶(ALT)和谷草转氨酶(AST)大于30 IU/L,女性大于19 IU/L 为升 高。采用卡方检验和多因素Logistic 回归分析,探讨一般特征和咖啡消费频率 与ALT 和AST 升高的风险。结果:咖啡摄入频率<1,1 和>=2 次/天的人群高 ALT 的发生率分别为27.4%、27.8%和26.9%,高AST 的发生率分别为 32.5%、33.1%和26.7%。咖啡摄入频率>=2 次/天的人群比<1 次/的人群调整 OR 显著降低(ALT:aOR=0.86,95% CI =0.79-0.94;AST:aOR=0.83,95% CI=0.76-0.91)。亚组分析显示,高危人群整体以及病毒性肝炎组和肥胖组中 咖啡摄入频率>=2 次/天的人群ALT 升高的风险低。敏感性分析显示:减少咖 啡消费频率与肝酶升高的风险增加相关,但在女性和当前吸烟者中并未发现 咖啡消费量与ALT 升高之间的关系。结论:韩国成人较高的咖啡消费量与氨 基转移酶浓度升高的风险较低有关。.

  16. Recurrent truncating mutations in alanine-glyoxylate aminotransferase gene in two South Indian families with primary hyperoxaluria type 1 causing later onset end-stage kidney disease

    PubMed Central

    Dutta, A. K.; Paulose, B. K.; Danda, S.; Alexander, S.; Tamilarasi, V.; Omprakash, S.

    2016-01-01

    Primary hyperoxaluria type 1 is an autosomal recessive inborn error of metabolism due to liver-specific peroxisomal enzyme alanine-glyoxylate transaminase deficiency. Here, we describe two unrelated patients who were diagnosed to have primary hyperoxaluria. Homozygous c.445_452delGTGCTGCT (p.L151Nfs*14) (Transcript ID: ENST00000307503; human genome assembly GRCh38.p2) (HGMD ID CD073567) mutation was detected in both the patients and the parents were found to be heterozygous carriers. Our patients developed end-stage renal disease at 23 years and 35 years of age. However, in the largest series published from OxalEurope cohort, the median age of end-stage renal disease for null mutations carriers was 9.9 years, which is much earlier than our cases. Our patients had slower progressions as compared to three unrelated patients from North India and Pakistan, who had homozygous c.302T>C (p.L101P) (HGMD ID CM093792) mutation in exon 2. Further, patients need to be studied to find out if c.445_452delGTGCTGCT mutation represents a founder mutation in Southern India. PMID:27512303

  17. Characterization of Potential Antimicrobial Targets in Bacillus spp. II. Branched-Chain Aminotransferase and Methionine Regeneration in B. cereus and B. anthracis

    DTIC Science & Technology

    2002-09-01

    chaine ramifi~e, encod6 par le g~ ne ybgE, catalysait la formation de la m~thionine. Cependant, le B. subtilis nWest pas un mod~le particuli~rement exact...programme TTCP. Puisque le B. cereus et le B. anthracis sont de la m~me esp~ce et qu’ils ne contiennent des diffdrences que dans le contenu plasmide...l’existence de donn~es signifiantes Sur les g6nomes font que le B. cereus est un organisme non pathog ~nique beaucoup plus appropri6 A la bicochimie du B

  18. A low-pungency S3212 genotype of Capsicum frutescens caused by a mutation in the putative aminotransferase (p-AMT) gene.

    PubMed

    Park, Young-Jun; Nishikawa, Tomotaro; Minami, Mineo; Nemoto, Kazuhiro; Iwasaki, Tomohiro; Matsushima, Kenichi

    2015-12-01

    The purpose of this study was to identify the genetic mechanism underlying capsinoid biosynthesis in S3212, a low-pungency genotype of Capsicum frutescens. Screening of C. frutescens accessions for capsaicinoid and capsiate contents by high-performance liquid chromatography revealed that low-pungency S3212 contained high levels of capsiate but no capsaicin. Comparison of DNA coding sequences of pungent (T1 and Bird Eye) and low-pungency (S3212) genotypes uncovered a significant 12-bp deletion mutation in exon 7 of the p-AMT gene of S3212. In addition, p-AMT gene transcript levels in placental tissue were positively correlated with the degree of pungency. S3212, the low-pungency genotype, exhibited no significant p-AMT transcript levels, whereas T1, one of the pungent genotypes, displayed high transcript levels of this gene. We therefore conclude that the deletion mutation in the p-AMT gene is related to the loss of pungency in placental tissue and has given rise to the low-pungency S3212 C. frutescens genotype. C. frutescens S3212 represents a good natural source of capsinoids. Finally, our basic characterization of the uncovered p-AMT gene mutation should contribute to future studies of capsinoid biosynthesis in Capsicum.

  19. Hepatic (Liver) Function Panel

    MedlinePlus

    ... related side effects. The hepatic function panel evaluates: Alanine aminotransferase (ALT). This enzyme, found in the liver, ... MORE ON THIS TOPIC Mononucleosis Hepatitis Blood Test: Alanine Aminotransferase (ALT, or SGPT) Blood Test: Aspartate Aminotransferase ( ...

  20. 78 FR 59696 - Leased Asset Energy and GHG Reporting Interpretive Guidance

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-09-27

    ... Performance Green Buildings (MG), Office of Government-Wide Policy, GSA, at 202-501-0762 or via email at kinga.... Dated: September 23, 2013. Kevin Kampschroer, Federal Director, Office of Federal High Performance Green Buildings, Office of Government-wide Policy. BILLING CODE 6820-14-P...

  1. 32 CFR Appendix B to Part 172 - Disposition of Amounts Collected From Successful Bidders

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... DEFENSE CONTRACTING DISPOSITION OF PROCEEDS FROM DOD SALES OF SURPLUS PERSONAL PROPERTY Pt. 172, App. B... from the bidder, a check shall be drawn on the suspense account and forwarded to GSA at the following... sale _ F3875 _ F3875. Upon receipt of the entire amount due from the bidder, a check shall be drawn...

  2. 32 CFR Appendix B to Part 172 - Disposition of Amounts Collected From Successful Bidders

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... DEFENSE CONTRACTING DISPOSITION OF PROCEEDS FROM DOD SALES OF SURPLUS PERSONAL PROPERTY Pt. 172, App. B... from the bidder, a check shall be drawn on the suspense account and forwarded to GSA at the following... sale _ F3875 _ F3875. Upon receipt of the entire amount due from the bidder, a check shall be drawn...

  3. 32 CFR Appendix B to Part 172 - Disposition of Amounts Collected From Successful Bidders

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... DEFENSE CONTRACTING DISPOSITION OF PROCEEDS FROM DOD SALES OF SURPLUS PERSONAL PROPERTY Pt. 172, App. B... from the bidder, a check shall be drawn on the suspense account and forwarded to GSA at the following... sale _ F3875 _ F3875. Upon receipt of the entire amount due from the bidder, a check shall be drawn...

  4. Shaping Democratic Identities and Building Citizenship Skills through Student Activism: Mexico's First Gay-Straight Alliance

    ERIC Educational Resources Information Center

    MacGillivray, Ian K.

    2005-01-01

    In democratic societies schools play a large role in helping students learn the values and skills necessary for adult participation in a free and open society. This article is a case study of a group of students who started a Gay-Straight Alliance (GSA) at a private American school in Mexico City. The students' struggle to form and keep their GSA,…

  5. Expanding the Meaning of Social Education: What the Social Studies Can Learn from Gay Straight Alliances

    ERIC Educational Resources Information Center

    Mayo, J. B., Jr.

    2013-01-01

    The author explores significant points of intersection between foundational tenets of the social studies and the lessons learned by students in the Gay Straight Alliance (GSA) at one Midwestern high school. He suggests new ways social studies researchers and teachers might conceptualize the ideas and themes promoted in GSAs and apply them directly…

  6. 40 CFR 799.5075 - Drinking water contaminants subject to testing.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... count. (13) Clinical biochemical determinations. Certain clinical biochemistry determinations on blood... aminotransferase, serum aspartate aminotransferase, gamma glutamyl transpeptidase, urea nitrogen, albumin, blood... investigation of observed effects. (14) Histopathology. Histopathology of the lungs of all animals shall...

  7. PSAT1 — EDRN Public Portal

    Cancer.gov

    From NCBI Gene: This gene encodes a member of the class-V pyridoxal-phosphate-dependent aminotransferase family. The encoded protein is a phosphoserine aminotransferase and decreased expression may be associated with schizophrenia. Mutations in this gene are also associated with phosphoserine aminotransferase deficiency. Alternative splicing results in multiple transcript variants. Pseudogenes of this gene have been defined on chromosomes 1, 3, and 8. [provided by RefSeq, Jul 2013

  8. Baseline hematology and clinical chemistry results from captive-raised trumpeter swans

    USGS Publications Warehouse

    Olsen, G.H.; Rininger, D.L.; Ets, M.K.; Sladen, William J. L.; Rees, Eileen C.; Earnst, Susan L.; Coulson, John C.

    2002-01-01

    Results from hematology and clinical chemistry tests are presented for healthy captive-raised Trumpeter Swans (Cygnus buccinator) to help establish baseline data. Blood samples were obtained from 14 cygnets between the ages of three to four and seven to eight months that were the subjects of a study to teach migration routes to swans. Males and females differed significantly in asparatate aminotransferase, alanine aminotransferase and total protein. Age categories differed significantly in hematocrit, white blood cell counts, alkaline phosphatase, aspar-rate aminotransferase, glucose, cholesterol and uric acid. There were no significant differences among age categories in values of alanine aminotransferase, calcium, triglycerides and total protein.

  9. Influence of Asymptomatic Pneumonia on the Response to Hemorrhage and Resuscitation in Swine

    DTIC Science & Technology

    2010-01-01

    dehydrogenase (LDH), aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatine kinase (CK), amylase and lactate (Vitros Chemistry System...CK. Creatinine increased at 15 min in both groups and remained elevated throughout the study. Mean total protein, amylase and ALT decreased similarly

  10. Toxicological evaluation of the hydro-alcohol extract of the dry leaves of Peumus boldus and boldine in rats.

    PubMed

    Almeida, E R; Melo, A M; Xavier, H

    2000-03-01

    The hydro-alcohol extract of the dry leaves of Peumus boldus and boldine, showed abortive and teratogenic action and changes in the blood levels of bilirubin, cholesterol, glucose, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and urea in rats. The long term administration of the extract and boldine did not cause histological modification during a period of 90 days.

  11. Genome Sequence of Klebsiella pneumoniae YZUSK-4, a Bacterium Proposed as a Starter Culture for Fermented Meat Products.

    PubMed

    Yu, Hai; Yin, Yongqi; Xu, Lin; Yan, Ming; Fang, Weiming; Ge, Qingfeng

    2015-07-23

    Klebsiella pneumoniae strain YZUSK-4, isolated from Chinese RuGao ham, is an efficient branched-chain aminotransferase-producing bacterium that can be used widely in fermented meat products to enhance flavor. The draft genome sequence of strain YZUSK-4 may provide useful genetic information on branched-chain amino acid aminotransferase production and branched-chain amino acid metabolism.

  12. Genome Sequence of Klebsiella pneumoniae YZUSK-4, a Bacterium Proposed as a Starter Culture for Fermented Meat Products

    PubMed Central

    Yu, Hai; Yin, Yongqi; Yan, Ming; Fang, Weiming; Ge, Qingfeng

    2015-01-01

    Klebsiella pneumoniae strain YZUSK-4, isolated from Chinese RuGao ham, is an efficient branched-chain aminotransferase-producing bacterium that can be used widely in fermented meat products to enhance flavor. The draft genome sequence of strain YZUSK-4 may provide useful genetic information on branched-chain amino acid aminotransferase production and branched-chain amino acid metabolism. PMID:26205853

  13. Safety of red ginseng oil for single oral administration in Sprague–Dawley rats

    PubMed Central

    Bak, Min-Ji; Kim, Kyu-Bong; Jun, Mira; Jeong, Woo-Sik

    2013-01-01

    The single oral administration of red ginseng oil (5000 mg/kg) to Sprague–Dawley rats induced no changes in behavioral patterns, clinical signs, and body weight, and hepatotoxicity parameters such as aspartate aminotransferase and alanine aminotransferase for 14 d. Therefore, these results suggest that the red ginseng oil is safe and nontoxic acutely. PMID:24558315

  14. Identification of the enzymatic basis for. delta. -aminolevulinic acid auxotrophy in a hemA mutant of escherichia coli

    SciTech Connect

    Avissar, Y.J.; Beale, S.I. )

    1989-06-01

    The hemA mutation of Escherichia coli K-12 confers a requirement for {delta}-aminolevulinic acid (ALA). Cell extract prepared from the hemA strain SASX41B was incapable of producing ALA from either glutamate or glutamyl-tRNA, whereas extract of the hem{sup +} strain HB101 formed colorimetrically detectable amounts of ALA and transferred label from 1-({sup 14}C)glutamate and 3,4-({sup 3}H)glutamyl-tRNA to ALA. Extracts of both strains converted glutamate-1-semialdehyde to ALA and were capable of aminoacylating tRNA{sup Glu}. Glutamyl-tRNA formed by extracts of both strains could be converted to ALA by the extract of hem{sup +} cells. The extract of hemA cells did not convert glutamyl-tRNA formed by either strain to ALA. However, the hemA cell extract, when supplemented in vitro with glutamyl-tRNA dehydrogenase isolated from Chlorella vulgaris cells, formed about as much ALA as did the unsupplemented hem{sup +} cell extract. We conclude from these observations that the enzyme activity that is lacking in the ALA auxotrophic strain carrying the hemA mutation is that of glutamyl-tRNA dehydrogenase.

  15. Suspected fusariomycotoxicosis in sandhill cranes (Grus canadensis): clinical and pathological findings.

    USGS Publications Warehouse

    Roffe, Thomas J.; Stroud, Richard K.; Windingstad, Ronald M.

    1989-01-01

    In 1985 and 1986, large-scale natural die-offs of sandhill cranes in Texas were attributed to fusariomycotoxicosis. These birds demonstrated a progressive loss of motor control to the neck, wings, and legs. Based on necropsy and/or histopathology of 31 cranes, the most common lesions involved skeletal muscle and included hemorrhages, granulomatous myositis, thrombosis, and vascular degeneration. Serum chemistry results revealed that levels of creatinine kinase, aspartate aminotransferase, and alanine aminotransferase were above published normals. However, only alanine aminotransferase was higher in clinically affected cranes than in normal cranes collected from the same area.

  16. Optimization of an Isolated Perfused Rainbow Trout Liver Model: Clearance Studies with 7-Ethoxycoumarin

    EPA Science Inventory

    Isolated trout livers were perfused using methods designed to preserve tissue viability and function. Liver performance was evaluated by measuring O2 consumption (VO2), vascular resistance, K+ leakage, glucose flux, lactate flux, alanine aminotransferase (ALT) leakage, and meta...

  17. Metabolism of 7-ethyoxycoumarin by Isolated Perfused Rainbow Trout Livers

    EPA Science Inventory

    Isolated trout livers were perfused using methods designed to preserve tissue viability and function. Liver performance was evaluated by measuring O2 consumption, vascular resistance, K+ leakage, glucose flux, lactate flux, alanine aminotransferase leakage, and metabolic clearanc...

  18. Blood Test: Comprehensive Metabolic Panel (CMP)

    MedlinePlus

    ... a decrease in kidney function. Alkaline phosphatase (ALP) , alanine amino transferase (ALT) , aspartate amino transferase (AST) , and ... Test: Glucose Kidneys and Urinary Tract Blood Test: Alanine Aminotransferase (ALT, or SGPT) Blood Test: Basic Metabolic ...

  19. Effects of dietary supplementation with vitamin C and vitamin E and their combination on growth performance, some biochemical parameters, and oxidative stress induced by copper toxicity in broilers.

    PubMed

    Cinar, Miyase; Yildirim, Ebru; Yigit, A Arzu; Yalcinkaya, Ilkay; Duru, Ozkan; Kisa, Uçler; Atmaca, Nurgul

    2014-05-01

    This study investigated effects of dietary supplementation with vitamin C, vitamin E on performance, biochemical parameters, and oxidative stress induced by copper toxicity in broilers. A total of 240, 1-day-old, broilers were assigned to eight groups with three replicates of 10 chicks each. The groups were fed on the following diets: control (basal diet), vitamin C (250 mg/kg diet), vitamin E (250 mg/kg diet), vitamin C + vitamin E (250 mg/kg + 250 mg/kg diet), and copper (300 mg/kg diet) alone or in combination with the corresponding vitamins. At the 6th week, the body weights of broilers were decreased in copper, copper + vitamin E, and copper + vitamin C + vitamin E groups compared to control. The feed conversion ratio was poor in copper group. Plasma aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase activities, iron, copper concentrations, and erythrocyte malondialdehyde were increased; plasma vitamin A and C concentrations and erythrocyte superoxide dismutase were decreased in copper group compared to control. Glutathione peroxidase, vitamin C, and iron levels were increased; aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and copper levels were decreased in copper + vitamin C group, while superoxide dismutase, glutathione peroxidase, and vitamin E concentrations were increased; aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase were decreased in copper with vitamin E group compared to copper group. The vitamin C concentrations were increased; copper, uric acid, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and malondialdehyde were decreased in copper + vitamin C + vitamin E group compared to copper group. To conclude, copper caused oxidative stress in broilers. The combination of vitamin C and vitamin E addition might alleviate the harmful effects of copper as demonstrated by decreased lipid peroxidation and hepatic enzymes.

  20. Antiviral Activity of a Small-Molecule Inhibitor of Filovirus Infection

    DTIC Science & Technology

    2010-05-01

    alanine aminotransferase (ALT), albumin, alkaline phosphatases (ALP), amylase , aspartate aminotransferase (AST), calcium, creatinine, -glutamyltransferase...types, the CC50 was calculated to be equivalent to or above 50 M (not shown). Thus, the results generated from these cell- FIG. 1. Chemical structure of...single 1-mg/kg dose by inducing a massive alpha interferon (IFN-) response in EBOV-infected cells (4, 5). Interestingly, 3-deazaneplanocin A induced a

  1. Liver Function Tests Following Open Cardiac Surgery

    PubMed Central

    Sabzi, Feridoun; Faraji, Reza

    2015-01-01

    Introduction: The cardiopulmonary bypass may have multiple systemic effects on the body organs as liver. This prospective study was planned to explore further the incidence and significance of this change. Methods: Two hundred patients with coronary artery bypass grafting (CABG), were randomly selected for the study. Total and indirect bilirubin, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase were measured preoperatively and at 24, 48 and 72 hours, following coronary artery bypass grafting. Postoperative value of the liver function tests with respect to hypothermia or hypotension were compared by one way analysis of variance for repeated measure and compared with t test. Patient’s characteristics with bilirubin value (≤1.5 mg or >1.5 mg) were compared with t test. Results: A significant increase of total bilirubin, aspartate aminotransferase, and alkaline phosphatase were noted in the third postoperative day. Significant relation was seen between hypotension and alkaline phosphatase, and aspartate aminotransferase change but hypothermia had not affected alanine aminotransferase, total bilirubin and indirect bilirubin change. Pump time, alanine aminotransferase in third postoperative day and direct bilirubin in first and second day of postoperative period had significant relation with pre and post-operative bilirubin change. Conclusion: Transient but not permanent alterations of hepatic enzymes after coronary artery bypass grafting presumably attributed to the decreased hepatic flow, hypoxia, or pump-induced inflammation. PMID:26191391

  2. A point mutation in Euglene gracilis chloroplast tRNA{sup Glu} uncouples protein and chlorophyll biosynthesis

    SciTech Connect

    Stange-Thomann, N.; Thomann, H.U.; Lloyd, A.J.; Soell, D.; Lyman, H.

    1994-08-16

    The universal precursor of tetrapyrrole pigments (e.g., chlorophylls and hemes) is 5-aminolevulinic acid (ALA), which in Euglena gracilis chloroplasts is derived via the two-step C{sub 5} pathway from glutamate charged to tRNA{sup Glu}. The first enzyme in this pathway, Glu-tRNA reductase (GluTR) catalyzes the reduction of glutamyl-tRNA{sup Glu} (Glu-tRNA) to glutamate 1-semialdehyde (GSA) with the release of the uncharged tRNA{sup Glu}. The second enzyme, GSA-2, 1-aminomutase, converts GSA to ALA. tRNA{sup Glu} is a specific cofactor for the NADPH-dependent reduction by GluTR, an enzyme that recognizes the tRNA in a sequence-specific manner. This RNA is the normal tRNA{sup Glu}, a dual-function molecule participating both in protein and in ALA and, hence, chlorophyll biosynthesis. A chlorophyll-deficient mutant of E. gracilis (Y{sub 9}ZNaIL) does not synthesize ALA from glutamate, although it contains GluTR and GSA-2,1-aminomutase activity. The tRNA{sup Glu} isolated from the mutant can still be acrylated with glutamate in vitro and in vitro. Furthermore, it supports chloroplast protein synthesis; however, it is a poor substrate for GluTR. Sequence analysis of the tRNA and of its gene revealed a C56 {yields} U mutation in the resulting gene product. C56 is therefore an important identity element for GluTR. Thus, a point mutation in the T loop of tRNA uncouples protein from chlorophyll biosynthesis.

  3. Cell organelles from crassulacean-acid-metabolism (CAM) plants : I. Enzymes in isolated peroxisomes.

    PubMed

    Herbert, M; Burkhard, C; Schnarrenberger, C

    1978-01-01

    Cell organelles were isolated from the CAM plants Crassula lycopodioides Lam., Bryophyllum calycinum Salisb. and Sedum rubrotinctum R.T. Clausen by isopycnic centrifugation in sucrose gradients. The inclusion of 2.5% Ficoll in the grinding medium proved to be essential for a satisfactory separation of cell organelles during the subsequent centrifugation. Peroxisomes, mitochondria, and whole and broken chloroplasts were at least partially resolved as judged by marker-enzyme-activity profiles. The isolated peroxisomes contained activities of glycollate oxidase, catalase, hydroxypyruvate reductase, glycine aminotransferase, serine-glyoxylate aminotransferase, and aspartate aminotransferase, comparable to activities found in spinach (Spinacia oleracea L.) leaf peroxisomes. In contrast to spinach, however, only little, if any, particulate malate dehydrogenase activity could be attributed to isolated peroxisomes of the three CAM plants.

  4. Assessment of liver function among nickel-plating workers in Egypt.

    PubMed

    El-Shafei, H M

    2011-06-01

    Currently no reports are available from Egypt regarding occupational exposure to nickel and its effects on the liver. The aim of this study was to assess the liver function of workers occupationally exposed to nickel. Standard liver function tests were applied to blood samples from 25 nickel-plating workers in Damietta, Egypt and 30 administrative workers as a reference group. Levels of urine nickel, measured by inductively coupling plasma-emission spectroscopy, were significantly higher in nickel-exposed workers compared with the reference group. The levels of alanine aminotransferase and aspartate aminotransferase were significantly higher in nickel-exposed workers. The level of serum albumin was significantly negatively correlated and the levels of serum aminotransferases, and serum gamma-glutamyl-transpeptidase were significantly positively correlated with urine nickel levels. Liver function is compromised in nickel-plating workers compared with non-exposed administrative workers.

  5. [Effect of space flight on the Kosmos-1129 biosatellite on enzyme activity of the rat liver].

    PubMed

    Nemeth, S; Tigranian, R A

    1983-01-01

    After the 18.5 day Cosmos-1129 flight the activity of 7 glucocorticoid-stimulated enzymes of the rat liver was measured. Immediately postflight the activity of tyrosine aminotransferase, tryptophan pyrolase and serine dehydrogenase increased. These enzymes rapidly (within several hours) react to increased glucocorticoids. The activity of aspartate and alanine aminotransferases also increased. These enzymes require many days of a continuous effect of glucocorticoids. The glycogen concentration in the rat liver also grew. At R + 6 the activity of tryptophan pyrolase and serine dehydrogenase decreased and that of the other enzymes returned to normal. The immobilization stress applied postflight led to an increased activity of tyrosine aminotransferase and tryptophan pyrolase. This study gives evidence that after space flight rats are in an acute stress state, evidently, produced by the biosatellite recovery.

  6. Asparagine Metabolic Pathways in Arabidopsis.

    PubMed

    Gaufichon, Laure; Rothstein, Steven J; Suzuki, Akira

    2016-04-01

    Inorganic nitrogen in the form of ammonium is assimilated into asparagine via multiple steps involving glutamine synthetase (GS), glutamate synthase (GOGAT), aspartate aminotransferase (AspAT) and asparagine synthetase (AS) in Arabidopsis. The asparagine amide group is liberated by the reaction catalyzed by asparaginase (ASPG) and also the amino group of asparagine is released by asparagine aminotransferase (AsnAT) for use in the biosynthesis of amino acids. Asparagine plays a primary role in nitrogen recycling, storage and transport in developing and germinating seeds, as well as in vegetative and senescence organs. A small multigene family encodes isoenzymes of each step of asparagine metabolism in Arabidopsis, except for asparagine aminotransferase encoded by a single gene. The aim of this study is to highlight the structure of the genes and encoded enzyme proteins involved in asparagine metabolic pathways; the regulation and role of different isogenes; and kinetic and physiological properties of encoded enzymes in different tissues and developmental stages.

  7. Effect of short-term exposure to three chemicals on the blood chemistry of the pinfish lagodon rhomboides

    SciTech Connect

    Folmar, L.C.; Bonomelli, S.; Moody, T.; Gibson, J.

    1993-01-01

    Injections of 3 ml/kg CCl4 caused significant elevations in the serum enzymes alanine aminotransferase (ALT), aspartate aminotransferase (AST), creatine kinase (CK) and lactate dehydrogenase (LD-L). Serum lipids and total protein were significantly lower, while serum glucose was significantly greater. Serum protein electrophoresis showed disassociation of albumin. Seawater species appear more tolerant of nitrite intoxication than freshwater species. Concentrations of fenthion as high as 30% of the 48-hr LC50 did not inhibit serum cholinesterase or alter serum chemistry.

  8. Acute metabolic effects of ammonia on the enzymes of glutamate metabolism in isolated astroglial cells.

    PubMed

    Subbalakshmi, G Y; Murthy, C R

    1983-01-01

    Enzymes of glutamate metabolism were studied in the astrocytes isolated from rats injected with a large dose of ammonium acetate and compared with those isolated from controls. The activities of glutamate dehydrogenase (GDH) and glutaminase decreased while those of glutamine synthetase (GS) and aspartate aminotransferase (AAT) increased both in convulsive and comatose states. The activity of alanine aminotransferase (A1AT) increased only in convulsive state. The results suggested that glutamate required for the formation of glutamine in astrocytes might have its origin in nerve endings and the depletion of citric acid cycle intermediates might occur in nerve endings at least in acute ammonia toxicity.

  9. Hepatoprotective effect of vitamin B12 on dimethylnitrosamine-induced liver injury.

    PubMed

    Isoda, Katsuhiro; Kagaya, Noritaka; Akamatsu, Soichiro; Hayashi, Shinji; Tamesada, Makoto; Watanabe, Aiko; Kobayashi, Masakazu; Tagawa, Yoh-ichi; Kondoh, Masuo; Kawase, Masaya; Yagi, Kiyohito

    2008-02-01

    Vitamin B(12) contains a cobalt complex and accumulates at high levels in the liver. Vitamin B(12) was examined for its hepatoprotective effect on dimethylnitrosamine-induced liver injury in mice. Vitamin B(12) decreased the blood levels of aspartate aminotransferase and alanine aminotransferase, and clearly inhibited the overaccumulation of collagen fibrils. Reverse transcription-polymerase chain reaction (RT-PCR) analysis of the liver showed that the gene expression of alpha-smooth muscle actin and heat-shock protein 47, which are markers of fibrosis, were suppressed by vitamin B(12) administration. Our findings indicate that vitamin B(12) could be an effective hepatoprotective agent.

  10. Prosopis farcta beans increase HDL cholesterol and decrease LDL cholesterol in ostriches (Struthio camelus).

    PubMed

    Omidi, Arash; Ansari nik, Hossein; Ghazaghi, Mahmood

    2013-02-01

    Ten blue-neck male ostriches (Struthio camelus) were fed Prosopis farcta beans throughout a 30-day experiment. Blood samples were collected from ostriches on days 0 and 30 to measure levels of high-density lipoprotein (HDL) cholesterol, low-density lipoprotein (LDL) cholesterol, triglyceride, total serum protein, albumin, globulin, cholesterol, calcium, inorganic phosphorus, the activity of aspartate aminotransferase, alanine aminotransferase, and γ-glutamyl transferase (γ-GT). From days 0 to 30, HDL cholesterol, total protein, and globulins levels increased significantly whereas LDL cholesterol, inorganic phosphorus, and γ-GT activity decreased significantly.

  11. Plasma chemistry references values in psittaciformes.

    PubMed

    Lumeij, J T; Overduin, L M

    1990-04-01

    Reference values for 17 plasma chemical variables in African greys. Amazons, cockatoos and macaws were established for use in avian clinical practice. The inner limits are given for the percentiles P(2.5) and P(97.5) with a probability of 90%. The following variables were studied: urea, creatinine, uric acid, urea/uric acid ratio, osmolality, sodium, potassium, calcium, glucose, aspartate aminotransferase, alanine aminotransferase, gamma glutamyltransferase, lactate dehydrogenase, creatine kinase, bile acids, total protein, albumin/globulin ratio. Differences between methods used and values found in this study and those reported previously are discussed.

  12. Autoimmune Hepatitis Associated with Immune Thrombocytopenic Purpura

    PubMed Central

    Ito, Akihiro; Yoshizawa, Kaname; Fujimori, Kazuya; Morita, Susumu; Shigeno, Takashi; Maejima, Toshitaka

    2017-01-01

    Although autoimmune hepatitis (AIH) is frequently complicated with chronic thyroiditis or other autoimmune disorders, reports on its association with immune thrombocytopenic purpura (ITP) are scarce. We herein describe a case of AIH associated with ITP. A 75-year-old Japanese woman was admitted to our hospital due to increased aminotransferase levels and severe thrombocytopenia. Elevated serum immunoglobulin G (IgG) was detected, and tests for platelet-associated IgG and anti-nuclear antibody were positive. Following the diagnosis of AIH-associated ITP, prednisolone treatment of 0.6 mg/kg/day resulted in a decrease in the aminotransferase levels and an increased platelet count. PMID:28090042

  13. A series of cases of rhabdomyolysis after ingestion of Tricholoma equestre

    PubMed Central

    Gabija, Laubner; Gabija, Mikulevičienė

    2016-01-01

    Tricholoma equestre (hereinafter – T. equestre) is a common edible fungus that is considered to be toxic under certain conditions. Here, we report four cases of acute poisoning caused by T. equestre, including one lethal outcome in Lithuania between 2004 and 2013. In the severe case, fatigue, nausea without vomiting and muscle pain, profuse sweating without fever, and respiratory insufficiency occurred. Laboratory tests showed an elevation of creatine kinase (CK), aspartate aminotransferase (AST), and alanine aminotransferase (ALT). Although clinical findings and laboratory tests support evidence of rhabdomyolysis, no renal insufficiency was observed. Significance of T. equestre in cardiac changes is feasible but remains unclear. PMID:28356809

  14. Effect of the aqueous extract of Syzygium cumini on carbon tetrachloride-induced hepatotoxicity in rats.

    PubMed

    Moresco, Rafael Noal; Sperotto, Rita Leal; Bernardi, Anie Schiavo; Cardoso, Ricardo França; Gomes, Patrícia

    2007-08-01

    The aim of this study was to evaluate the effect of the aqueous Syzygium cumini leaf extract, given either as a single dose or by 7 days of pretreatment, on hepatotoxicity induced by carbon tetrachloride in rats. Blood samples obtained after treatments were measured for aspartate aminotransferase (AST) and alanine aminotransferase (ALT). A significant increase in the AST and ALT activities occurred after carbon tetrachloride administration alone, which was significantly lowered by preadministration with the aqueous extract of Syzygium cumini, but not by a single dose. This suggests that the extract may be useful for liver protection but needs to be given over a significant period and prior to liver injury.

  15. Enzyme activities in plasma, liver, and kidney of black ducks and mallards

    USGS Publications Warehouse

    Franson, J. Christian

    1982-01-01

    Activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST), creatine phosphokinase (CPK), alkaline phosphatase (ALP), and lactate dehydrogenase (LDH) were measured in plasma, liver, and kidney, and gamma-glutamyl transferase (GGT) was measured in liver and kidney of black ducks (Anas rubripes). Activities of ALT, AST, GGT, and ornithine carbamyl transferase (OCT) were assayed in plasma, liver, and kidney of game-farm mallards (Anas platyrhynchos). Appreciable OCT and AST activity occurred in both liver and kidney. Activities of ALT, CPK, ALP and GGT were higher in kidney, while LDH was higher in liver, GGT was detected in plasma from one of four mallards.

  16. Radiology case of the month. Nausea, vomiting, and diarrhea in a patient with hepatitis C and acquired immunodeficiency syndrome (AIDS). Diffuse, severe gastric-wall thickening, consistent with edema.

    PubMed

    Mabry, Christian; Hutchings, John; Sanders, Charles; Neitzschman, Harold

    2012-01-01

    The patient is a 42-year-old male with a past medical history of HIV/AIDS (his most recent CD4 count, four months before admission, was 19) and hepatitis C who presented to the Emergency Department complaining of one week of persistent nausea, vomiting, and diarrhea. His admit labs were as follows: hemoglobin of 11.8, hematocrit of 35, total protein of 6.0, albumin of 1.6, total bilirubin of 2.3, aspartate aminotransferase (AST) of 141, alkaline phosphatase (ALP) of 146, and alanine aminotransferase (ALT) of 31. Computed tomography (CT) images of the abdomen and pelvis with contrast were obtained (Figures 1 - 4).

  17. Infrequent Clinical Assessment of Chronic Hepatitis B Patients in United States General Healthcare Settings.

    PubMed

    Spradling, Philip R; Xing, Jian; Rupp, Loralee B; Moorman, Anne C; Gordon, Stuart C; Teshale, Eyasu T; Lu, Mei; Boscarino, Joseph A; Trinacty, Connie M; Schmidt, Mark A; Holmberg, Scott D

    2016-11-01

    Among 2338 chronic hepatitis B patients followed during 2006-2013 in the Chronic Hepatitis Cohort Study, 78% had ≥1 alanine aminotransferase and 37% had ≥1 hepatitis B virus DNA level assessed annually. Among cirrhotic patients, 46% never had hepatic imaging. Patients in this cohort were insufficiently monitored for disease activity and hepatocellular carcinoma.

  18. Precursors for ornithine and citrulline synthesis in neonatal pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Citrulline (CIT) is an amino acid synthesized by gut and utilized for the synthesis of the conditionally essential amino acid arginine (ARG). In turn, the immediate precursor for CIT synthesis, ornithine (ORN), can originate from proline (PRO) and glutamine (GLN) via ornithine aminotransferase (OAT,...

  19. [Effect of low-intensity 900 MHz frequency electromagnetic radiation on rat liver and blood serum enzyme activities].

    PubMed

    Nersesova, L S; Petrosian, M S; Gazariants, M G; Mkrtchian, Z S; Meliksetian, G O; Pogosian, L G; Akopian, Zh I

    2014-01-01

    The comparative analysis of the rat liver and blood serum creatine kinase, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase and purine nucleoside phosphorylase post-radiation activity levels after a total two-hour long single and fractional exposure of the animals to low-intensity 900 MHz frequency electromagnetic field showed that the most sensitive enzymes to the both schedules of radiation are the liver creatine kinase, as well as the blood serum creatine kinase and alkaline phosphatase. According to the comparative analysis of the dynamics of changes in the activity level of the liver and blood serum creatine kinase, alanine aminotransferase, aspartate aminotransferase and purine nucleoside phosphorylase, both single and fractional radiation schedules do not affect the permeability of a hepatocyte cell membrane, but rather cause changes in their energetic metabolism. The correlation analysis of the post-radiation activity level changes of the investigated enzymes did not reveal a clear relationship between them. The dynamics of post-radiation changes in the activity of investigated enzyme levels following a single and short-term fractional schedules of radiation did not differ essentially.

  20. Hepatoprotective Effect of Cuscuta campestris Yunck. Whole Plant on Carbon Tetrachloride Induced Chronic Liver Injury in Mice

    PubMed Central

    Peng, Wen-Huang; Chen, Yi-Wen; Lee, Meng-Shiou; Chang, Wen-Te; Tsai, Jen-Chieh; Lin, Ying-Chih; Lin, Ming-Kuem

    2016-01-01

    Cuscuta seeds and whole plant have been used to nourish the liver and kidney. This study was aimed to investigate the hepatoprotective activity of the ethanol extract of Cuscuta campestris Yunck. whole plant (CCEtOH). The hepatoprotective effect of CCEtOH (20, 100 and 500 mg/kg) was evaluated on carbon tetrachloride (CCl4)-induced chronic liver injury. Serum alanine aminotransferase, aspartate aminotransferase, triglyceride and cholesterol were measured and the fibrosis was histologically examined. CCEtOH exhibited a significant inhibition of the increase of serum alanine aminotransferase, aspartate aminotransferase, triglyceride and cholesterol. Histological analyses showed that fibrosis of liver induced by CCl4 were significantly reduced by CCEtOH. In addition, 20, 100 and 500 mg/kg of the extract decreased the level of malondialdehyde (MDA) and enhanced the activities of anti-oxidative enzymes including superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GRd) in the liver. We demonstrate that the hepatoprotective mechanisms of CCEtOH were likely to be associated to the decrease in MDA level by increasing the activities of antioxidant enzymes such as SOD, GPx and GRd. In addition, our findings provide evidence that C. campestris Yunck. whole plant possesses a hepatoprotective activity to ameliorate chronic liver injury. PMID:27941627

  1. Schisandra chinensis fruit modulates the gut microbiota composition in association with metabolic markers in obese women: a randomized, double-blind placebo-controlled study.

    PubMed

    Song, Mi-young; Wang, Jing-hua; Eom, Taewoong; Kim, Hojun

    2015-08-01

    Schisandra chinensis fruit (SCF) is known to have beneficial effects on metabolic diseases, including obesity, and to affect gut microbiota in in vivo studies. However, in human research, there have been a few studies in terms of its clinical roles in lipid metabolism and modulation of gut microbiota. A double-blind, placebo-controlled study with 28 obese women with SCF or placebo was conducted for 12 weeks. Anthropometry and blood and fecal sampling were performed before and after treatment. Analysis of the gut microbiota in feces was performed using denaturing gradient gel electrophoresis and quantitative polymerase chain reaction. Although the values did not differ significantly between the 2 groups, the SCF group tended to show a greater decrease in waist circumference, fat mass, fasting blood glucose, triglycerides, aspartate aminotransferase, and alanine aminotransferase than the placebo group. Clustering of the denaturing gradient gel electrophoresis fingerprints for total bacteria before and after treatment indicated more separate clustering in SCF group than placebo. In correlation analysis, Bacteroides and Bacteroidetes (both increased by SCF) showed significant negative correlation with fat mass, aspartate aminotransferase, and/or alanine aminotransferase, respectively. Ruminococcus (decreased by SCF) showed negative correlation with high-density lipoprotein cholesterol and fasting blood glucose. In conclusion, administration of SCF for 12 weeks resulted in modulation of the gut microbiota composition in Korean obese women, and significant correlations with some bacterial genera and metabolic parameters were noted. However, in general, SCF was not sufficient to induce significant changes in obesity-related parameters compared with placebo.

  2. Removing a bottleneck in the Bacillus subtilis biotin pathway: bioA utilizes lysine rather than S-adenosylmethionine as the amino donor in the KAPA-to-DAPA reaction.

    PubMed

    Van Arsdell, Scott W; Perkins, John B; Yocum, R Rogers; Luan, Linda; Howitt, C Linda; Chatterjee, Nilu Prasad; Pero, Janice G

    2005-07-05

    In biotin biosynthesis, DAPA aminotransferase encoded by the bioA gene catalyzes the formation of the intermediate 7,8-diaminopelargonic acid (DAPA) from 7-keto-8-aminopelargonic acid (KAPA). DAPA aminotransferases from Escherichia coli, Serratia marcescens, and Bacillus sphaericus use S-adenosylmethionine (SAM) as the amino donor. Our observation that SAM is not an amino donor for B. subtilis DAPA aminotransferase led to a search for an alternative amino donor for this enzyme. Testing of 26 possible amino acids in a cell-free extract assay revealed that only l-lysine was able to dramatically stimulate the in vitro conversion of KAPA to DAPA by the B. subtilis DAPA aminotransferase. The K(m) for lysine and KAPA was estimated to be between 2 and 25 mM, which is significantly higher than the K(m) of purified E. coli BioA for SAM (0.15 mM). This higher requirement for lysine resulted in accumulation of KAPA during fermentation of B. subtilis biotin producing strains. However, this pathway bottleneck could be relieved by either addition of exogenous lysine to the medium or by introduction of lysine deregulated mutations into the production strains.

  3. Remnants of an ancient pathway to L-phenylalanine and L-tyrosine in enteric bacteria: Evolutionary implications and biotechnological impact. [Escherichia coli; Salmonella typhimurium; Neurospora crassa

    SciTech Connect

    Bonner, C.A.; Fischer, R.S.; Ahmad, S.; Jensen, R.A. )

    1990-12-01

    The pathway construction for biosynthesis of aromatic amino acids in Escherichia coli is atypical of the phylogenetic subdivision of gram-negative bacteria to which it belongs. Related organisms possess second pathways to phenylalanine and tyrosine which depend upon the expression of a monofunctional chorismate mutase (CM-F) and cyclohexadienyl dehydratase (CDT). Some enteric bacteria, unlike E. coli, possess either CM-F or CDT. These essentially cryptic remnants of an ancestral pathway can be a latent source of biochemical potential under certain conditions. As one example of advantageous biochemical potential, the presence of CM-F in Salmonella typhimurium increases the capacity for prephenate accumulation in a tyrA auxotroph. We report the finding that a significant fraction of the latter prephenate is transaminated to L-arogenate. The tyrA19 mutant is now the organism of choice for isolation of L-arogenate, uncomplicated by the presence of other cyclohexadienyl products coaccumulated by a Neurospora crassa mutant that had previously served as the prime biological source of L-arogenate. Prephenate aminotransferase activity was not conferred by a discrete enzyme, but rather was found to be synonymous with the combined activities of aspartate aminotransferase (aspC), aromatic aminotransferase (tyrB), and branched-chain aminotransferase (ilvE).

  4. 75 FR 69353 - Isoxaben; Pesticide Tolerances

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-11-12

    ... the following activities: Crop production (NAICS code 111). Animal production (NAICS code 112). Food... histopathology and increased blood alkaline phosphatase and alanine aminotransferase activities at high doses. In... hypertrophy and induction of hepatic microsomal enzymes. Increased incidence and severity of nephropathy...

  5. Biochemical stress response in freshwater fish Channa punctatus induced by aqueous extracts of Euphorbia tirucalli plant.

    PubMed

    Tiwari, Sudhanshu; Singh, Ajay

    2006-06-01

    Piscicidal activities of aqueous extracts of Euphorbia tirucalli were very well established, but their ultimate mode of action on fish metabolism was not yet known. Exposure of fishes over 24h or 96h to sub-lethal doses (40% and 80% of LC(50)) of aqueous extract of E. tirucalli stem-bark and latex, significantly (P<0.05) altered the level of total protein, total free amino acids, nucleic acids, glycogen, pyruvate, lactate and activity of protease, alanine aminotransferase, aspartate aminotransferase, acetylcholinesterase and cytochrome oxidase enzyme in liver and muscle tissues of freshwater fish Channa punctatus. The alterations in all these biochemical parameters were significantly (P<0.05) time- and dose-dependent. After 7d of withdrawal of treatment of 80% of LC(50) of E. tirucalli extracts shows that there was a partial recovery in the levels of glycogen but nearly complete recovery in total protein, total free amino acids, pyruvate, lactate, nucleic acids level and activity of protease, aspartate aminotransferase, alanine aminotransferase, acetylcholinesterase and cytochrome oxidase enzyme in both the tissues of fish. Thus aqueous extracts of E. tirucalli adversely affect respiratory pathway of fish and cause energy crisis during stress by suppressing ATP production. The reversibility of the action of the aqueous extracts would be an additional advantage in their use.

  6. Protection Against Microcystin-LR-Induced Hepatoxicity by Silymarin: Biochemistry, Histopathology and Lethality

    DTIC Science & Technology

    1990-04-04

    wild artichoke (jilybus sdrinum L. Gaertn), completely abolihed the lethal effects, pathological changes, and ,34nificantly decreased the levels of...aminotransferase, and lactate dehydrogenase. Pretreatment of either rats or mice with a single dose of silymarin, a flavonotignane isolated from the wild artichoke

  7. Clinical and pathological characteristics of hepatotoxicity associated with occupational exposure to dimethylformamide

    SciTech Connect

    Redlich, C.A.; West, A.B.; Fleming, L.; True, L.D.; Cullen, M.R.; Riely, C.A. )

    1990-09-01

    The clinical characteristics, laboratory results, and liver biopsy findings of seven workers with toxic liver injury associated with exposure to several solvents, including substantial levels of the widely used solvent dimethylformamide, are presented. Three patients had short exposure (less than 3 months), four long exposure (greater than 1 year). Among those with brief exposure, symptoms included anorexia, abdominal pain, and disulfiram-type reaction. Aminotransferases were markedly elevated with the ratio of alanine aminotransferase to aspartate aminotransferase always greater than 1. Liver biopsy showed focal hepatocellular necrosis and microvesicular steatosis with prominence of smooth endoplasmic reticulum, complex lysosomes, and pleomorphic mitochondria with crystalline inclusions. Among workers with long exposure, symptoms were minimal and enzyme elevations modest. Biopsies showed macrovesicular steatosis, pleomorphic mitochondria without crystalloids, and prominent smooth endoplasmic reticulum, but no evidence of persisting acute injury or fibrosis. Abnormal aminotransferases in both groups may persist for months after removal from exposure, but progression to cirrhosis in continually exposed workers was not observed. We conclude that exposure of these workers to solvents, chiefly dimethylformamide, may result in two variants of toxic liver injury with subtle clinical, laboratory, and morphological features. This may be readily overlooked if occupational history and biopsy histology are not carefully evaluated.

  8. Chronic hepatitis E virus infection in a pediatric female liver transplant recipient.

    PubMed

    Passos-Castilho, Ana Maria; Porta, Gilda; Miura, Irene K; Pugliese, Renata P S; Danesi, Vera L B; Porta, Adriana; Guimarães, Teresa; Seda, João; Antunes, Eduardo; Granato, Celso F H

    2014-12-01

    We describe a case of chronic hepatitis E virus (HEV) infection in a 13-year-old female liver transplant recipient with recurrent increased aminotransferase levels and acute cellular rejection. This finding demonstrates that chronic HEV infection can occur and should be further investigated in immunocompromised patients in Latin America.

  9. Antioxidative Role of Hatikana (Leea macrophylla Roxb.) Partially Improves the Hepatic Damage Induced by CCl4 in Wistar Albino Rats.

    PubMed

    Akhter, Samina; Rahman, Md Atiar; Aklima, Jannatul; Hasan, Md Rakibul; Chowdhury, J M Kamirul Hasan

    2015-01-01

    This research investigated the protective role of Leea macrophylla extract on CCl4-induced acute liver injury in rats. Different fractions of Leea macrophylla (Roxb.) crude extract were subjected to analysis for antioxidative effects. Rats were randomly divided into four groups as normal control, hepatic control, and reference control (silymarin) group and treatment group. Evaluations were made for the effects of the fractions on serum enzymes and biochemical parameters of CCl4-induced albino rat. Histopathological screening was also performed to evaluate the changes of liver tissue before and after treatment. Different fractions of Leea macrophylla showed very potent 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging effect, FeCl3 reducing effect, superoxide scavenging effect, and iron chelating effect. Carbon tetrachloride induction increased the level of serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) and other biochemical parameters such as lipid profiles, total protein, and CK-MB. In contrast, treatment of Leea macrophylla reduced the serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) activities as well as biochemical parameters activities. L. macrophylla partially restored the lipid profiles, total protein, and CK-MB. Histopathology showed the treated liver towards restoration. Results evidenced that L. macrophylla can be prospective source of hepatic management in liver injury.

  10. Elevated liver enzymes associated with dronedarone for atrial fibrillation

    PubMed Central

    2013-01-01

    A 51-year-old male with documented atrial fibrillation who was taking dronedarone 400 mg twice daily for approximately 3 months returned to the cardiologist for an ablation procedure. Baseline liver enzymes were within normal range prior to starting the medication and increased after the 3 months of therapy. Aspartate aminotransferase increased from 31 IU/L to 98 IU/L, and alanine aminotransferase increased from 21 IU/L to 101 IU/L. Two and a half months after discontinuation of the medication, liver enzymes normalized (aspartate aminotransferase: 30 IU/L and alanine aminotransferase: 25 IU/L). The Food and Drug Administration has now alerted health-care professionals of the potential for liver injury based upon post-marketing surveillance. The chronological course of elevated liver enzymes noted in our patient is suggestive of a dronedarone-induced problem. Clinicians should have a heightened awareness of the potential for liver enzyme elevation and injury with dronedarone and should monitor enzymes periodically, especially within the first 6 months of use. PMID:27489632

  11. Development of Toxicity Data for Munition Compounds to Support Toxicity Reference Value Derivations for Wildlife

    DTIC Science & Technology

    2010-06-01

    lacked righting reflex. Anesthesia prior to euthanasia was achieved via carbon dioxide exposure, and euthanasia was completed through decapitation and...10 concentrations of compound up to 5000 milligrams per kilograms (mg/kg) and observed daily. They were fed between 10–20 flightless fruit flies...the following clinical chemistries: albumin (ALB), alkaline phosphatase (ALP), aspartate aminotransferase (AST), calcium (Ca), cholesterol (CHOL

  12. Identification and Characterization of Distinct Apoptotic Pathways in Cancer Cells Activated in Response to Treatment with Different Anti-Cancer Agents

    DTIC Science & Technology

    1999-07-01

    fuller understanding of the role of apoptosis in real tumor progression such as breast cancer and will lead to the developing new strategies for anti...382. Fagan, R.J., Lazaris-Karatzas, A., Sonenberg, N., and Rozen , R. Translational control of ornithine aminotransferase. J Biol Chem 266, 16518-16523

  13. Antialcoholic liver activity of whey fermented by Lactobacillus casei isolated from koumiss.

    PubMed

    Zhao, Z W; Pan, D D; Wu, Z; Sun, Y Y; Guo, Y X; Zeng, X Q

    2014-07-01

    Whey fermented liquid (WFL) was studied for its hepatoprotective effects by using chronic alcohol-induced mice. Whey fermented liquid, prepared by inoculating whey with 4% (vol/vol) Lactobacillus casei and then incubating at 41°C for 8h, was used to orally treat alcohol-induced mice at 3 dosages for 5 wk. Ethanol consumption significantly reduced the activity of superoxide dismutase and glutathione peroxidase, while lowering glutathione content and increasing levels of aspartate aminotransferase, alanine aminotransferase, total triglyceride, malondialdehyde, and cytochrome P450 2E1. Treatment with WFL significantly attenuated the increased levels of alanine aminotransferase, aspartate aminotransferase, triglyceride, and cytochrome P450 2E1, while decreasing superoxide dismutase, glutathione peroxidase, malondialdehyde, and glutathione levels. Pathological changes in the livers of mice who had ingested alcohol were improved by the administration of WFL. These results suggest that WFL may exert a protective effect against alcoholic liver disease by increasing antioxidant activity, which supports the use of WFL as an antialcoholic liver disease treatment.

  14. Monosodium glutamate-induced damage in liver and kidney: a morphological and biochemical approach.

    PubMed

    Ortiz, G G; Bitzer-Quintero, O K; Zárate, C Beas; Rodríguez-Reynoso, S; Larios-Arceo, F; Velázquez-Brizuela, I E; Pacheco-Moisés, F; Rosales-Corral, S A

    2006-02-01

    It has been demonstrated that high concentrations of monosodium glutamate in the central nervous system induce neuronal necrosis and damage in retina and circumventricular organs. In this model, the monosodium glutamate is used to induce an epileptic state; one that requires highly concentrated doses. The purpose of this study was to evaluate the toxic effects of the monosodium glutamate in liver and kidney after an intra-peritoneal injection. For the experiment, we used 192 Wistar rats to carry out the following assessments: a) the quantification of the enzymes alanine aminotransferase and aspartate aminotransferase, b) the quantification of the lipid peroxidation products and c) the morphological evaluation of the liver and kidney. During the experiment, all of these assessments were carried out at 0, 15, 30 and 45 min after the intra-peritoneal injection. In the rats that received monosodium glutamate, we observed increments in the concentration of alanine aminotransferase and aspartate aminotransferase at 30 and 45 min. Also, an increment of the lipid peroxidation products, in kidney, was exhibited at 15, 30 and 45 min while in liver it was observed at 30 and 45 min. Degenerative changes were observed (edema-degeneration-necrosis) at 15, 30 and 45 min.

  15. Effect of Polysaccharide from Cordyceps militaris (Ascomycetes) on Physical Fatigue Induced by Forced Swimming.

    PubMed

    Xu, Yan-Feng

    2016-01-01

    Cordyceps militaris is the one of the most important medicinal mushrooms, widely used in East Asian countries. Polysaccharide is considered to be the principal active component in C. militaris and has a wide range of biological and pharmacological properties. This study was undertaken to investigate the effect of polysaccharide from C. militaris (PCM) on physical fatigue induced in animals through a forced swimming test. The mice were divided into 4 groups receiving 28 days' treatment with drinking water (exercise control) or low-, medium-, and high-dose PCM (40, 80, and 160 mg/kg/day, respectively). After 28 days, the mice were subjected to the forced swimming test; the exhaustive swimming time was measured and fatigue-related biochemical parameters, including serum lactic acid, urea nitrogen, creatine kinase, alanine aminotransferase, aspartate aminotransferase, superoxide dismutase, glutathi- one peroxidase, catalase, malondialdehyde, liver glycogen, and muscle glycogen, were analyzed. The results showed that PCM could significantly prolong the exhaustive swimming time of mice; decrease concentrations of serum lactic acid, urea nitrogen, creatine kinase, aspartate aminotransferase, alanine aminotransferase, and malondialdehyde; and increase liver and muscle glycogen contents and the concentrations of serum superoxide dismutase, glutathione per- oxidase, and catalase. The data suggest that PCM has an antifatigue effect, and it might become a new functional food or medicine for fatigue resistance.

  16. Investigation of the possible protective role of gallic acid on paraoxanase and arylesterase activities in livers of rats with acute alcohol intoxication.

    PubMed

    Kartkaya, Kazim; Oğlakçi, Ayşegül; Şentürk, Hakan; Bayramoğlu, Gökhan; Canbek, Mediha; Kanbak, Güngör

    2013-04-01

    Gallic acid, a polyphenyl class natural product from gallnut and green tea, is known to be antioxidant, anti-inflammatory and radical scavenger. In this study, we aimed to investigate the possible protective effects of gallic acid on paraoxonase and arylesterase activities in liver exposed to acute alcohol intoxication. Paraoxonase and arylesterase activities in liver tissue and serum aspartate aminotransferase, alanine aminotransferase and lactate dehydrogenase levels were measured. Histological investigations were also made. In our study, we observed a significant increase of serum alanine aminotransferase, aspartate aminotransferase and lactate dehydrogenase activities, which are indicators of liver damage after acute ethanol consumption. Gallic acid therapy has significantly reduced the increase in these biomarkers, indicating a possible hepatoprotective effect of gallic acid. Ethanol consumption caused a significant decrease in liver paraoxonase activity (P < 0.001). Gallic acid treatment partly restored this decreased paraoxonase activity, which resulted from ethanol administration. A gallic acid dose of 100 mg/kg was observed as highest restoring effect for paraoxonase activity (P < 0.05). The activity of arylesterase was decreased in the ethanol group as compared with the control group, but this was not significant. However, 50 mg/kg of gallic acid treatment restored the loss of this activity due to ethanol exposure (P < 0.001). We observed that gallic acid ameliorates the liver damage caused by excessive alcohol consumption in a dose-dependent way. Our results in this study showed that gallic acid might have a protective effect against alcoholic liver disease.

  17. Heat shock proteins 27 and 70 contribute to the protection of Schisandrin B against d-galactosamine-induced liver injury in mice.

    PubMed

    Gao, Zhiying; Zhang, Jishun; Li, Libo; Shen, Longqing; Li, Qingyi; Zou, Yu; Du, Xiaohui; Zhao, Zhibo

    2016-04-01

    Schisandrin B is a hepatoprotective component isolated from a traditional Chinese herb, Schisandra chinensis (Turcz.) Baill. This study determined the effect of Schisandrin B on d-galactosamine -induced liver injury and the role of heat shock proteins 27 and 70 against liver injury in mice. Acute liver injury was induced by intraperitoneal injection of d-galactosamine to mice, and Schisandrin B was given orally. The protein and gene expression of heat shock proteins 27 and 70 were detected by western blot and real-time quantitative polymerase chain reaction, respectively. Liver tissues were subjected to histological evaluation, and the activities of alanine aminotransferase and aspartate aminotransferase in the serum were measured. Pretreatment of Schisandrin B significantly attenuated d-galactosamine-induced liver injury in mice. This result was evidenced by improved alteration of histopathological hepatic necrosis and reduced alanine aminotransferase and aspartate aminotransferase activities in the serum. The hepatoprotective effect was accompanied with overexpression of heat shock proteins 27 and 70 both at the protein and mRNA levels. However, the aforementioned actions of Schisandrin B were all markedly suppressed by the heat shock protein inhibitor quercetin. Heat shock proteins 27 and 70 were involved in the protective effect of Schisandrin B against d-galactosamine-induced liver injury in mice.

  18. Crystal structures of Mycobacterium tuberculosis HspAT and ArAT reveal structural basis of their distinct substrate specificities

    PubMed Central

    Nasir, Nazia; Anant, Avishek; Vyas, Rajan; Biswal, Bichitra Kumar

    2016-01-01

    Aminotransferases of subfamily Iβ, which include histidinol phosphate aminotransferases (HspATs) and aromatic amino acid aminotransferases (ArATs), are structurally similar but possess distinct substrate specificities. This study, encompassing structural and biochemical characterisation of HspAT and ArAT from Mycobacterium tuberculosis demonstrates that the residues lining the substrate binding pocket and N-terminal lid are the primary determinants of their substrate specificities. In mHspAT, hydrophilic residues in the substrate binding pocket and N-terminal lid allow the entry and binding of its preferential substrate, Hsp. On the other hand, the hydrophobic nature of both the substrate binding pocket and the N-terminal lid of mArAT is responsible for the discrimination of a polar substrate such as Hsp, while facilitating the binding of Phe and other aromatic residues such as Tyr and Trp. In addition, the present study delineates the ligand induced conformational rearrangements, providing insights into the plasticity of aminotransferases. Furthermore, the study also demonstrates that the adventitiously bound ligand 2-(N-morpholino)ethanesulfonic acid (MES) is indeed a specific inhibitor of HspAT. These results suggest that previously untapped morpholine-ring scaffold compounds could be explored for the design of new anti-TB agents. PMID:26738801

  19. Detection of TTV in peripheral blood cells from patients with altered ALT and AST levels.

    PubMed

    de Oliveira, Jaqueline Carvalho; Nasser, Thiago Franco; Oda, Julie Massayo Maeda; Aoki, Mateus Nóbrega; Carneiro, Juliana Laino do Val; Barbosa, Décio Sabbatini; Reiche, Edna Maria Vissoci; Watanabe, Maria Angelica Ehara

    2008-04-01

    This work analyzes the prevalence of TTV DNA in peripheral blood cells from patients with hepatic alterations and healthy blood donors and measures levels of sodium, potassium, urea, creatinine, phosphatase alkaline, total and direct bilirubin, gamma glutamyl transferase (GGT), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in certain randomly selected patients. DNA samples from 111 individuals were evaluated. They were divided into two groups, "A" (study) and "B" (control), including 54 patients with liver enzyme alterations (ALT/AST) presenting non-B-non-C hepatitis and 57 blood donors, respectively. TTV DNA was determined by nested PCR. Certain products of the second-round PCR were sequenced. Serum biochemical assay was performed and disclosed TTV in 31.48% (17/54) of patients in group A and 5.26% (3/57) in the control group B. TTV prevalence was significantly higher in patients with liver disease than in healthy donors. In group A, sodium, potassium, urea, creatinine, phosphatase alkaline, total and direct bilirubin, gamma glutamyl transferase (GGT), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were analyzed in certain randomly selected patients and no significant difference in biochemical levels (p>0.05) was found when TTV infected and noninfected individuals were compared. Knowledge related to TTV has rapidly increased, but many fundamental aspects remain unclear. This led us to question the role of TTV and doubt remains as to whether or not it is just a commensal virus. Further studies are necessary to confirm and extend these findings.

  20. Ringer's malate solution protects against the multiple organ injury and dysfunction caused by hemorrhagic shock in rats.

    PubMed

    Dai, Zhong-Liang; Wu, Jing; Meng, Chen; Zeng, Fang; Yang, Yan; Yao, Shang-Long

    2012-08-01

    Malic acid, in the form of its anion malate, is a key intermediate in the major biochemical energy-producing cycle known as the citric acid or Krebs cycle. In this study, the authors investigated the protective effect of a novel crystalloid solution of Ringer's malate following fluid resuscitation of hemorrhagic shock using a rat model. Under general anesthesia, Sprague-Dawley male rats were subjected to 60 min of hemorrhagic shock (40 mmHg for 60 min) followed by crystalloid resuscitation. Groups were as follows: (1) sham shock, (2) normal saline, (3) Ringer's lactate, and (4) Ringer's malate. The experiment was terminated at 4 h after resuscitation. Mean arterial blood pressure (MAP) and blood biophysical parameters were monitored during the experiment. The alanine aminotransferase, aspartate aminotransferase, urea, creatinine, superoxide dismutase, and malondialdehyde levels in plasma were detected. The intestine, liver, lung, and renal histopathology were measured. It was found that Ringer's malate could increase MAP immediately and maintain MAP for a long time. Ringer's malate could reduce the level of alanine aminotransferase, aspartate aminotransferase, urea, and creatinine. At the same time, the activity of superoxide dismutase was increased, and the level of malondialdehyde was decreased. Histopathology indicated that Ringer's malate can protect against the multiple organ injury caused by hemorrhagic shock in rats. Ringer's malate prevented circulatory failure and alleviated multiple organ dysfunction syndrome in animals with hemorrhagic shock. The study suggests that Ringer's malate solution could be a potential novel therapeutic agent for fluid resuscitation.

  1. Serum chemistry reference values in adult Japanese quail (Coturnix coturnix japonica) including sex-related differences.

    PubMed

    Scholtz, N; Halle, I; Flachowsky, G; Sauerwein, H

    2009-06-01

    Serum chemistry reference values may provide useful information about the physical condition of individuals, making them a useful tool in differentiating normal and healthy animals from abnormal or diseased states. For Japanese quail that are used for producing eggs and meat for human consumption and also as laboratory animals, we aimed to extend the available array of reference values and to compare 16-wk-old adult male versus female birds. In the present study, clinical chemistry data (albumin, total protein, glucose, uric acid, cholesterol, bilirubin, cholinesterase, creatinine, triglycerides, alanine aminotransferase, aspartate aminotransferase, and gamma-glutamyltransferase) in blood serum from up to 125 male and 151 female Japanese quail were established. Statistical comparisons were made between male and female birds. Aspartate aminotransferase, alanine aminotransferase, glucose, cholinesterase, and bilirubin values were higher (P < 0.01) in males, whereas females had higher (P < 0.05) concentrations of albumin, total protein, gamma-glutamyltransferase, total cholesterol, and triglycerides. No significant sex-based differences were observed for creatinine and uric acid. The reference values provided are relevant in particular for the use of quail as laboratory animals when responses to specific treatments have to be monitored and appraised.

  2. Antioxidative Role of Hatikana (Leea macrophylla Roxb.) Partially Improves the Hepatic Damage Induced by CCl4 in Wistar Albino Rats

    PubMed Central

    Akhter, Samina; Rahman, Md. Atiar; Aklima, Jannatul; Hasan, Md. Rakibul; Hasan Chowdhury, J. M. Kamirul

    2015-01-01

    This research investigated the protective role of Leea macrophylla extract on CCl4-induced acute liver injury in rats. Different fractions of Leea macrophylla (Roxb.) crude extract were subjected to analysis for antioxidative effects. Rats were randomly divided into four groups as normal control, hepatic control, and reference control (silymarin) group and treatment group. Evaluations were made for the effects of the fractions on serum enzymes and biochemical parameters of CCl4-induced albino rat. Histopathological screening was also performed to evaluate the changes of liver tissue before and after treatment. Different fractions of Leea macrophylla showed very potent 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging effect, FeCl3 reducing effect, superoxide scavenging effect, and iron chelating effect. Carbon tetrachloride induction increased the level of serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) and other biochemical parameters such as lipid profiles, total protein, and CK-MB. In contrast, treatment of Leea macrophylla reduced the serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) activities as well as biochemical parameters activities. L. macrophylla partially restored the lipid profiles, total protein, and CK-MB. Histopathology showed the treated liver towards restoration. Results evidenced that L. macrophylla can be prospective source of hepatic management in liver injury. PMID:26221590

  3. CORRELATION BETWEEN RAINFALL PATTERNS AND THE WATER TABLE IN THEGENERAL SEPARATIONS AREA OF THE SAVANNAH RIVERSITE

    SciTech Connect

    Smith, C.

    2009-08-10

    The objective of the study was to evaluate rainfall and water table elevation data in search of a correlation that could be used to understand and predict water elevation changes. This information will be useful in placing screen zones for future monitoring wells and operations of groundwater treatment units. Fifteen wells in the General Separations Area (GSA) at Savannah River Site were evaluated from 1986 through 2001. The study revealed that the water table does respond to rainfall with minimal delay. (Water level information was available monthly, which restricted the ability to evaluate a shorter delay period.) Water elevations were found to be related to the cumulative sum (Q-Delta Sum) of the difference between the average rainfall for a specific month and the actual rainfall for that month, calculated from an arbitrary starting point. Water table elevations could also be correlated between wells, but using the right well for correlation was very important. The strongest correlation utilized a quadratic equation that takes into account the rainfall in a specific area and the rainfall from an adjacent area that contributes through a horizontal flow. Specific values vary from well to well as a result of geometry and underground variations. R2's for the best models ranged up to 0.96. The data in the report references only GSA wells but other wells (including confined water tables) on the site have been observed to return similar water level fluctuation patterns.

  4. Radionuclide inventories for the F- and H-area seepage basin groundwater plumes

    SciTech Connect

    Hiergesell, Robert A; Kubilius, Walter P.

    2016-05-01

    Within the General Separations Areas (GSA) at the Savannah River Site (SRS), significant inventories of radionuclides exist within two major groundwater contamination plumes that are emanating from the F- and H-Area seepage basins. These radionuclides are moving slowly with groundwater migration, albeit more slowly due to interaction with the soil and aquifer matrix material. The purpose of this investigation is to quantify the activity of radionuclides associated with the pore water component of the groundwater plumes. The scope of this effort included evaluation of all groundwater sample analyses obtained from the wells that have been established by the Environmental Compliance & Area Completion Projects (EC&ACP) Department at SRS to monitor groundwater contamination emanating from the F- and H-Area Seepage Basins. Using this data, generalized groundwater plume maps for the radionuclides that occur in elevated concentrations (Am-241, Cm-243/244, Cs-137, I-129, Ni-63, Ra-226/228, Sr-90, Tc-99, U-233/234, U-235 and U-238) were generated and utilized to calculate both the volume of contaminated groundwater and the representative concentration of each radionuclide associated with different plume concentration zones.

  5. Regulation of the hemA gene during 5-aminolevulinic acid formation in Pseudomonas aeruginosa.

    PubMed Central

    Hungerer, C; Troup, B; Römling, U; Jahn, D

    1995-01-01

    The general tetrapyrrole precursor 5-aminolevulinic acid is formed in bacteria via two different biosynthetic pathways. Members of the alpha group of the proteobacteria use 5-aminolevulinic acid synthase for the condensation of succinyl-coenzyme A and glycine, while other bacteria utilize a two-step pathway from aminoacylated tRNA(Glu). The tRNA-dependent pathway, involving the enzymes glutamyl-tRNA reductase (encoded by hemA) and glutamate-1-semialdehyde-2,1-aminomutase (encoded by hemL), was demonstrated to be used by Pseudomonas aeruginosa, Pseudomonas putida, Pseudomonas stutzeri, Comamonas testosteroni, Azotobacter vinelandii, and Acinetobacter calcoaceticus. To study the regulation of the pathway, the glutamyl-tRNA reductase gene (hemA) from P. aeruginosa was cloned by complementation of an Escherichia coli hemA mutant. The hemA gene was mapped to the SpeI A fragment and the DpnIL fragment of the P. aeruginosa chromosome corresponding to min 24.1 to 26.8. The cloned hemA gene, coding for a protein of 423 amino acids with a calculated molecular mass of 46,234 Da, forms an operon with the gene for protein release factor 1 (prf1). This translational factor mediates the termination of the protein chain at the ribosome at amber and ochre codons. Since the cloned hemA gene did not possess one of the appropriate stop codons, an autoregulatory mechanism such as that postulated for the enterobacterial system was ruled out. Three open reading frames of unknown function transcribed in the opposite direction to the hemA gene were found. hemM/orf1 and orf2 were found to be homologous to open reading frames located in the 5' region of enterobacterial hemA genes. Utilization of both transcription start sites was changed in a P. aeruginosa mutant missing the oxygen regulator Anr (Fnr analog), indicating the involvement of the transcription factor in hemA expression. DNA sequences homologous to one half of an Anr binding site were detected at one of the determined

  6. Determination of serum biochemical reference intervals for the Iberian lynx (Lynx pardinus).

    PubMed

    García, Ignacio; Napp, Sebastián; Zorrilla, Irene; Vargas, Astrid; Pastor, Josep; Muñoz, Alvaro; Martínez, Fernando

    2010-02-01

    Biochemical reference intervals were determined for 31 clinically healthy Iberian lynxes (Lynx pardinus) between 1 and 6 years of age. Thirteen of the lynxes were wild-caught and the other 18 were captive-reared animals. The samples were collected between November 2004 and December 2006. The influence of sex (males vs. females), age (juveniles vs. adults) and habitat condition (free-living vs. captive) on the biochemical analytes were evaluated. Serum albumin concentrations were significantly higher in females than in males, while creatine phosphokinase was higher in males. The levels of alkaline phosphatase and lactate dehydrogenase were higher in juvenile lynxes, while gamma glutamyl-transferase and creatinine values were higher in adults. Lynxes captured in the wild had higher concentrations of iron, calcium, alkaline phosphatase and creatinine, but lower aspartate aminotransferase and alanine aminotransferase than lynxes maintained in captivity. The results were generally comparable to commonly reported reference intervals for other lynx species, the domestic cat and other felid species.

  7. Comparison of blood chemistry values for samples collected from juvenile chinook salmon by three methods

    USGS Publications Warehouse

    Congleton, J.L.; LaVoie, W.J.

    2001-01-01

    Thirteen blood chemistry indices were compared for samples collected by three commonly used methods: caudal transection, heart puncture, and caudal vessel puncture. Apparent biases in blood chemistry values for samples obtained by caudal transection were consistent with dilution with tissue fluids: alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), creatine kinase (CK), triglyceride, and K+ were increased and Na+ and Cl- were decreased relative to values for samples obtained by caudal vessel puncture. Some enzyme activities (ALT, AST, LDH) and K+ concentrations were also greater in samples taken by heart puncture than in samples taken by caudal vessel puncture. Of the methods tested, caudal vessel puncture had the least effect on blood chemistry values and should be preferred for blood chemistry studies on juvenile salmonids.

  8. Levels of transaminases, alkaline phosphatase, and protein in tissues of Clarias gariepienus fingerlings exposed to sublethal concentrations of cadmium chloride.

    PubMed

    Velmurugan, Babu; Selvanayagam, Mariadoss; Cengiz, Elif I; Uysal, Ersin

    2008-12-01

    The freshwater fish, Clarias gariepienus fingerlings, were exposed to sublethal concentrations (1.7 and 3.4 mg/L) of cadmium chloride for 12 days. Aspartate aminotransferase (AAT), alanine aminotransferase (ALT), alkaline phosphatase (ALP), and total protein levels were assayed in the gill, brain, and muscle of the fish at regular intervals of 6 and 12 days. The activities of AAT, ALT, and ALP of the treated fishes increased significantly in all the tissues compared with the control fish. Protein level in all the tissues showed a significant decrease in comparison to unexposed controls throughout the experimental periods. These results revealed that cadmium chloride effects the intermediary metabolism of C. gariepienus fingerlings and that the assayed enzymes can work as good biomarkers of contamination.

  9. Alternations in the liver enzymatic activity of Common carp, Cyprinus carpio in response to parasites, Dactylogyrus spp. and Gyrodactylus spp.

    PubMed

    Rastiannasab, Abulhasan; Afsharmanesh, Shiva; Rahimi, Ruhollah; Sharifian, Iman

    2016-12-01

    The present study was carried out to investigate the effects of parasites, monogenea, Dactylogyrus spp. and Gyrodactylus spp. on some enzymatic and biochemical components of liver in healthy and infected common carp, Cyprinus carpio. For this purpose, 10 healthy and 10 infected fish were collected from farm. The blood samples were taken and after separation of serum, the values of Aspartate aminotransferase (AST), Alanine aminotransferase (ALT) enzymes activities as well as Creatinine and Urea were measured. Based on obtained results, the values of AST, ALT enzymes activities as well as Creatinine and Urea were higher in the infected fish compared to non-infected fish. In conclusion; our results reveals that infection with external parasites, Dactylogyrus spp. and Gyrodactylus spp. can causes some dysfunctions in liver and kidney of common carp.

  10. Evaluation of commercial enzyme reagent kits by use of a semiautomated chemistry analyzer.

    PubMed

    Beckala, H R; Agrell, J; Forsman, R W; Homburger, H A

    1979-08-01

    The overall performances of several enzyme reagent kits for alkaline phosphatase, creatine kinase, lactic dehydrogenase, and aspartate aminotransferase were evaluated using an ABA-100 Bichromatic Analyzer. Interassay precision using this instrument with commercial reagents compared well with published data for similar analyses performed at university hospitals and referral laboratories. Significantly poorer precision with lower limits of linearity was observed when reagents recommended for use at 30 C were used at 37 C. Significant differences in measured levels of creatine kinase, lactic dehydrogenase, and aspartate aminotransferase due to different lots of expendable cuvettes were found for elevated levels of these enzymes. All kit reagents met manufacturers' claims for stability; however, different absolute levels of lactic dehydrogenase were observed with one kit reagent on successive days. Slight hemolysis affected creatine kinase levels measured with some reagent kits significantly more than others.

  11. Comparison of two dry chemistry analyzers and a wet chemistry analyzer using canine serum.

    PubMed

    Lanevschi, Anne; Kramer, John W.

    1996-01-01

    Canine serum was used to compare seven chemistry analytes on two tabletop clinical dry chemistry analyzers, Boehringer's Reflotron and Kodak's Ektachem. Results were compared to those obtained on a wet chemistry reference analyzer, Roche Diagnostic's Cobas Mira. Analytes measured were urea nitrogen (BUN), creatinine, glucose, aspartate aminotransferase (AST), alanine aminotransferase (ALT), cholesterol and bilirubin. Nine to 12 canine sera with values in the low, normal, and high range were evaluated. The correlations were acceptable for all comparisons with correlation coefficients greater than 0.98 for all analytes. Regression analysis resulted in significant differences for both tabletop analyzers when compared to the reference analyzer for cholesterol and bilirubin, and for glucose and AST on the Kodak Ektachem. Differences appeared to result from proportional systematic error occurring at high analyte concentrations.

  12. The Protective Effects of Buzui on Acute Alcoholism in Mice

    PubMed Central

    Wen, Da-Chao; Gao, Shu-di; Hu, Xiao-yu; Yi, Cheng

    2016-01-01

    This study was designed to investigate the role of a traditional buzui recipe in anti-inebriation treatment. Buzui consists of Fructus Schisandrae Chinensis, Fructus Chebulae, Fructus Mume, Fructus Crataegi, Endothelium Corneum Gigeriae Galli, and Excrementum Bombycis. The buzui mixture was delivered by gavage, and ethanol was delivered subsequent to the final treatment. The effects of buzui on the righting reflex, inebriation rates, and the survival curve are depicted. Blood alcohol concentrations, alanine aminotransferase (ALT) levels, aspartate aminotransferase (AST) levels, and alkaline phosphatase (ALP) levels were recorded. The activities of alcohol dehydrogenase (ADH), aldehyde dehydrogenase (ALDH), and superoxide dismutase (SOD), as well as malonaldehyde (MDA) levels, were also measured. Our results demonstrated that a traditional buzui recipe showed significant effects on promoting wakefulness and the prevention of acute alcohol intoxication, accelerating the metabolism of alcohol in the liver and reducing the oxidative damage caused by acute alcoholism. PMID:26884793

  13. Haematological and biochemical alterations caused by epidural and intramuscular administration of xylazine hydrochloride in dromedary camels (Camelus dromedarius).

    PubMed

    Azari, Omid; Molaei, Mohammad Mahdi; Emadi, Ladan; Sakhaee, Ehsanollah; Sharifi, Hamid; Mehdizadeh, Sara

    2012-01-01

    This study was conducted in 16 healthy immature dromedary camels weighing 120-150 kg to evaluate and compare the effects of epidural and intramuscular injections of xylazine administered at 0.1 mg/kg and 0.2 mg/kg. Haematological parameters included haemoglobin, packed cell volume, total erythrocyte count and total leukocyte count. Biochemical parameters included alkaline phosphates, alanine aminotransferase, aspartate aminotransferase, blood urea nitrogen, creatinine and glucose. Parameters were measured at different intervals before (baseline) and after the administration of drugs. Our study showed that the effect of xylazine on haematological and biochemical parameters is dose-dependant and is also related to the route of administration. The low dose of xylazine administered using both intramuscular and epidural methods showed minimal effects, whereas high doses of the drug, especially when injected intramuscularly, caused greater changes in haematological and biochemical parameters.

  14. [Three sporadic cases of acute hepatitis E].

    PubMed

    Kim, Dong Han; Park, Hyeuk; Moon, Seung Won; Jeong, Jong Hyuk; Yang, Hyuk Seung; Kim, Do Hyun; Kim, Ho Dong

    2007-08-01

    Acute hepatitis E is an endemic disease, commonly reported in Indian subcontinent, China, Africa, Central America, and so forth. It is a self-limiting disease like other acute hepatitis except in pregnant patient. Although sporadic hepatitis E is noted all over the world, most of them are associated with travel history to HEV-endemic area. In Korea, Hepatitis E is rarely reported. Moreover, sporadic acute hepatitis E without travel history to HEV-endemic area is very rare. We experienced three sporadic cases of acute hepatitis E, without travel history. All of them presented acute hepatitis symptoms, elevated aminotransferase, and positive IgM HEV Ab. Symptoms and aminotransferase levels were normalized during hospitalization and IgM HEV Ab converted negative after 4-8 months. We report three sporadic cases of onset-acute hepatitis E without travel history to HEV-endemic area.

  15. Trimethoprim-sulfamethoxazole-induced Steven Johnson syndrome in an HIV-infected patient.

    PubMed

    Taqi, Syed Ahmed; Zaki, Syed Ahmed; Nilofer, Angadi Rajasab; Sami, Lateef Begum

    2012-01-01

    Trimethoprim-sulfamethoxazole (TMP/SMX) is a widely prescribed antimicrobial for the management of several uncomplicated infections. It is commonly used for the treatment and prophylaxis of Pneumocystis jirovecii pneumonia (PCP) in the HIV-infected population. The adverse reaction to TMP/SMX is more frequent and severe in HIV-infected patients as compared to the general population. Here, we report a case of Stevens-Johnson syndrome (SJS) secondary to TMP/SMX. The patient had a generalized cutaneous reaction with involvement of the eyes, oral cavity, and genitals. He had elevated hepatic alanine aminotransferase and aspartate aminotransferase enzyme. TMP/SMX therapy was stopped and supportive treatment was started. His condition improved after eight days of stopping TMP/SMX therapy.

  16. Possible health effects of liquefied petroleum gas on workers at filling and distribution stations of Gaza governorates.

    PubMed

    Sirdah, M M; Al Laham, N A; El Madhoun, R A

    2013-03-01

    Liquefied petroleum gas (LPG) is widely used in the Gaza Strip for domestic purposes, in agriculture and industry and, illegally, in cars. This study aimed to identify possible health effects on workers exposed to LPG in Gaza governorates. Data were collected by a questionnaire interview, and haematological and biochemical analyses of venous blood samples were made from 30 workers at filling and distribution stations and 30 apparently healthy controls. Statistically significant differences were found in all self-reported health-related complaints among LPG workers versus controls. LPG workers had significantly higher values of red blood cell counts, haemoglobin, haematocrit mean corpuscular haemoglobin and platelet counts. They also had significantly higher values of kidney function tests (urea, creatinine and uric acid) and liver function enzyme activities (aspartate aminotransferase and alanine aminotransferase). LPG workers at Gaza Strip petroleum stations are at higher risk for health-related symptoms and clinical abnormalities.

  17. Hematologic and plasma chemistry values in captive psittacine birds.

    PubMed

    Polo, F J; Peinado, V I; Viscor, G; Palomeque, J

    1998-01-01

    Reference values for some hematologic parameters in 19 species and plasma chemical values in 11 species of Psittacine birds, including cockatoos, parrots, amazons, macaws, conures, and lories, were established for use in veterinary medicine. The following parameters were studied: hematocrit, hemoglobin concentration, erythrocyte number, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, erythrocyte dimensions, leukocyte number and differential leukocyte count, glucose, urea, uric acid, cholesterol, triglycerides, creatinine, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, creatinine phosphokinase, lactic dehydrogenase, gamma glutamyl transpeptidase, total plasma protein, albumin, globulins, albumin-globulin ratio, sodium, potassium, calcium, magnesium, total phosphorus, chloride, and osmolality. Hematologically, the Psittacine is a very homogeneous avian group, with small differences between species. They are, however, different from other groups of birds.

  18. Hematologic and plasma biochemical reference values in Indian peafowl (Pavo cristatus).

    PubMed

    Samour, Jaime; Naldo, Jesus; Rahman, Habeeb; Sakkir, Mohammed

    2010-06-01

    Blood samples were collected from captive, adult, clinically normal Indian peafowl (Pavo cristatus) for hematologic and plasma biochemical analyses. Hematologic parameters investigated were total red blood cell count, hemoglobin, packed cell volume, fibrinogen, mean cell volume, mean cell hemoglobin, mean cell hemoglobin concentration, total white blood cell count, differential white blood cell count, and thrombocyte count. Plasma biochemical parameters investigated were alanine aminotransferase, alkaline phosphatase, amylase, aspartate aminotransferase, bile acids, total bilirubin, blood urea nitrogen, calcium, cholesterol, creatinine, creatine kinase, gamma glutamyltransferase, lactate dehydrogenase, glucose, iron, phosphorus, and uric acid, as well as plasma protein electrophoresis. Results were compared with values from studies done in houbara bustards (Chlamydotis undulata), kori bustards (Ardeotis kori), stone curlews (Burhinus oedicnemus), and taxonomically related species, including ring-necked pheasants (Phasianus colchicus), red-legged partridges (Alectoris rufa), Kashmir native fowl (Kashmirfavorella), and Bangladesh native, Fayoumi, and Assil fowl (Gallus domesticus).

  19. Effect of copper on liver and bone metabolism in malnutrition.

    PubMed

    Güler, A H; Sapan, N; Ediz, B; Genç, Z; Ozkan, K

    1994-01-01

    This study was planned to investigate the effects of copper (Cu) deficiency on liver and bone metabolism in malnourished children. Serum total calcium (Ca), inorganic phosphorus (P), Ca/P, Cu/Ca, Cu/P ratios and alkaline phosphatase (ALP) activity values were analyzed. Aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma glutamyltransferase (GGT) enzyme activities and the ALT/AST (De Ritis) ratio as well as their correlations with Cu were tested to determine liver function. The results of the study showed that Cu deficiency directly affects the organic matrix formation, and by the suppression of ALP activity, indirectly causes decalcification. In the liver, however, no direct effect of Cu deficiency was seen. Deterioration in liver function and Cu deficiency increased parallel with the severity of malnutrition. Thus we concluded that a correlation exists between Cu and the parameters that indicate liver function.

  20. Novel bead-based platform for direct detection of unlabelled nucleic acids through Single Nucleobase Labelling.

    PubMed

    Venkateswaran, Seshasailam; Luque-González, Maria Angélica; Tabraue-Chávez, Mavys; Fara, Mario Antonio; López-Longarela, Barbara; Cano-Cortes, Victoria; López-Delgado, Francisco Javier; Sánchez-Martín, Rosario María; Ilyine, Hugh; Bradley, Mark; Pernagallo, Salvatore; Díaz-Mochón, Juan José

    2016-12-01

    Over the last decade, circulating microRNAs have received attention as diagnostic and prognostic biomarkers. In particular, microRNA122 has been demonstrated to be an early and more sensitive indicator of drug-induced liver injury than the widely used biomarkers such as alanine aminotransferase and aspartate aminotransferase. Recently, microRNA122 has been used in vitro to assess the cellular toxicity of new drugs and as a biomarker for the development of a rapid test for drug overdose/liver damage. In this proof-of-concept study, we report a PCR-free and label-free detection method that has a limit of detection (3 standard deviations) of 15 fmoles of microRNA122, by integrating a dynamic chemical approach for "Single Nucleobase Labelling" with a bead-based platform (Luminex(®)) thereby, in principle, demonstrating the exciting prospect of rapid and accurate profiling of any microRNAs related to diseases and toxicology.

  1. Institutional experience with clofarabine and cytarabine in relapsed pediatric acute myeloid leukemia.

    PubMed

    Moreno, Lucas; Fernandez-Navarro, Jose Maria; Del Mar Andres, Maria; Bautista, Francisco; Tasso, Maria; Verdeguer, Amparo

    2012-01-01

    Cytarabine (1000 mg/m/d intravenous for 5 d) and clofarabine (40 mg/m/d intravenous for 5 d) were given every 28 days to 9 children with relapsed acute myeloid leukemia at our institution. Among 19 courses, there were 18 infectious episodes. Median hospitalization time was 13 days (7.7 to 30.5 d) per cycle. Hepatobiliary abnormalities included alanine aminotransferase/aspartate aminotransferase elevation and hyperbilirubinemia. Four patients achieved complete remission (one after an earlier allogeneic Haematopoietic Progenitor Cell Transplant). Four patients are alive disease free. In summary, a proportion of children responded and was able to receive allogeneic Haematopoietic Progenitor Cell Transplant. Side effects were tolerable, although hospitalization time was prolonged.

  2. Lophirones B and C Attenuate Acetaminophen-Induced Liver Damage in Mice: Studies on Hepatic, Oxidative Stress and Inflammatory Biomarkers.

    PubMed

    Ajiboye, Taofeek O

    2016-10-01

    Lophirones B and C are chalcone dimers with proven chemopreventive activity. This study evaluates the hepatoprotective effect lophirones B and C in acetaminophen-induced hepatic damage in mice using biomarkers of hepatocellular indices, oxidative stress, proinflammatory factors and lipid peroxidation. Oral administrations of lophirones B and C significantly (p < 0.05) attenuated acetaminophen-mediated alterations in serum alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, albumin and total bilirubin. Similarly, acetaminophen-mediated decrease in activities of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase and glucose 6- phosphate dehydrogenase were significantly attenuated in the liver of mice. Increased levels of conjugated dienes, lipid hydroperoxides, malondialdehyde, protein carbonyl and fragmented DNA were significantly lowered by lophirones B and C. Levels of tumour necrosis factor-α, interleukin-6 and 8 were significantly lowered in serum of acetaminophen treated mice by the chalcone dimers. Overall, results of this study show that lophirones B and C halted acetaminophen-mediated hepatotoxicity.

  3. Influence of coffee brew in metabolic syndrome and type 2 diabetes.

    PubMed

    Abrahão, Sheila Andrade; Pereira, Rosemary Gualberto Fonseca Alvarenga; de Sousa, Raimundo Vicente; Lima, Adriene Ribeiro; Crema, Gabriela Previatti; Barros, Bianca Sacramento

    2013-06-01

    This study aimed to evaluate the effect of coffee drinking on clinical markers of diabetes and metabolic syndrome in Zucker rats. Diabetic Zucker rats with metabolic syndrome and control Zucker rats were used for in vivo tests. The animals received daily doses of coffee drink by gavage for 30 days. After the treatment, the levels of glucose, triglycerides, total cholesterol and fractions, creatinine, uric acid, activity of aspartate aminotransferase and alanine aminotransferase were evaluated. Urea and creatinine levels were also analyzed in urine. By collaborating in the modulation of the metabolic syndrome and diabetes mellitus type 2, coffee drink helped in reducing serum glucose, total cholesterol and triglycerides. The results demonstrate that treatment with roasted coffee drink, because of its hypoglycemic and hypolipidemic effect, is efficient in the protection of animals with metabolic syndrome and diabetes mellitus type 2.

  4. [Serum enzyme alterations during diabetic ketoacidosis. Prospective study of the behaviour of AP, AsAT, AIAT, and GGPT in 24 cases (author's transl)].

    PubMed

    Pérez-Jiménez, F; Muñoz, R; Jiménez Perepérez, J; Jiménez Alonso, J; Grilo, A; Benito, P

    1981-09-25

    Because of the scarcity of data on the changes of serum enzymes during diabetic ketoacidosis, the authors have prospectively studied the alterations of gamma-glutamyl transpeptidase (GGTP), alkaline phosphatase (AP), aspartate aminotransferase (AsAT), and alanine aminotransferase (AIAT) in this metabolic disturbance. The most significant finding was the frequent increase of AP activity on admission, with a systematic fall of the serum levels within the first 24 hours (p less than 0.001). This phenomenon was negatively correlated with patients' age (p less than 0.05), and the same occurred with the bone isoenzyme of AP (p less than 0.01). The remaining enzymes studies were always normal, thus suggesting that any increase of the serum levels of GGTP, AsAT, and AIAT found in a patient with diabetic ketoacidosis must arise the suspicion of an associated disturbance.

  5. [Coexistence of Celiac Disease and autoimmune hepatitis case study and literature review].

    PubMed

    Tagle, Martín; Nolte, Cecilia; Luna, Eduardo; Scavino, Yolanda

    2006-01-01

    The case of a patient who was initially diagnosed with Systemic Lupus Erythematosus, with subsequent documentation of Celiac Disease histologically and serologically is reported. The patient presented elevation of the aminotransferases, upon detection of the Celiac Disease which was initially attributed to the underlying disease. However, despite the complete resolution of her articular symptoms with a gluten-free diet, the liver chemistry abnormalities persisted. This led to consider an autoimmune hepatitis as the cause which was documented with a liver biopsy three months after the diagnosis of the celiac disease and under a strict gluten-free diet. Treatment with prednisone and azathioprine was initiated with complete normalization of aminotransferase levels. We present the sequence of events with the results and a review of the literature.

  6. Hydroxychloroquine-induced toxic hepatitis in a patient with systemic lupus erythematosus: a case report.

    PubMed

    Abdel Galil, S M

    2015-05-01

    Increased serum level of liver enzymes is a common finding in patients with systemic lupus erythematosus (SLE). Hepatotoxic drugs, viral hepatitis and fatty liver are thought to be the main causes of hepatic lesion in these patients. Our aim was to determine the cause of strikingly elevated liver enzymes in a case with systemic lupus presenting with acute abdomen. Liver enzyme abnormality was defined as a 10-fold or greater increase in aspartate aminotransferase and alanine aminotransferase. Acute toxic hepatitis was diagnosed, which rapidly returned to normal after cessation of the suspected causative medication, hydroxychloroquine, and subsequent administration of mycophenolate mofetil. Elevated liver enzymes are a major concern and should be well investigated in SLE patients.

  7. Exertional myopathy in whooping cranes (Grus americana) with prognostic guidelines.

    PubMed

    Hanley, Christopher S; Thomas, Nancy J; Paul-Murphy, Joanne; Hartup, Barry K

    2005-09-01

    Exertional myopathy developed in three whooping cranes (Grus americana) secondary to routine capture, handling, and trauma. Presumptive diagnosis of exertional myopathy was based on history of recent capture or trauma, clinical signs, and elevation of aspartate aminotransferase, alanine aminotransferase, creatine kinase, lactate dehydrogenase, and serum potassium. Treatments were attempted in each case, but ultimately were not successful. Gross and microscopic lesions at necropsy confirmed the diagnosis in each case, with the leg musculature most severely affected. Guidelines for determining prognosis of exertional myopathy in cranes have been included based on the analysis of these cases and others in the literature. As treatment is largely unrewarding, prevention remains the key in controlling exertional myopathy. Identification of predisposing factors and proper handling, immobilization, and transportation techniques can help prevent development of exertional myopathy in cranes.

  8. CORAL: binary classifications (active/inactive) for Liver-Related Adverse Effects of Drugs.

    PubMed

    Toropov, Andrey A; Toropova, Alla P; Rasulev, Bakhtiyor F; Benfenati, Emilio; Gini, Giuseppina; Leszczynska, Danuta; Leszczynski, Jerzy

    2012-09-01

    Classification data related to the Liver-Related Adverse Effects of Drugs have been studied with the CORAL software (http://www.insilico.eu/coral). Two datasets which contain compounds with two serum enzyme markers of liver toxicity: alanine aminotransferase (ALT, n=187) and aspartate aminotransferase (AST, n=209) are analyzed. Statistical quality of the prediction for ALT activity is n=35, Sensitivity = 0.5556, Specificity = 0.8077, and Accuracy = 0.7429. In the case of AST activity the prediction is characterized by n=42, Sensitivity = 0.6875, Specificity = 0.7692, and Accuracy = 0.7381. A number of structural alerts which can be related to the studied activities are revealed. It is the first attempt to build up the classification QSAR model by means of the Monte Carlo technique based on representation of the molecular structure by SMILES using the CORAL software.

  9. Aromatic hydrocarbon pathology in fish following a large spill into the Nemadji River, Wisconsin, USA

    SciTech Connect

    Caldwell, C.A.

    1997-04-01

    On June 30, 1992, a train accident resulted in a rail car releasing 114,000 L of a complex mixture of aromatic hydrocarbons into the Nemadji River, a tributary of Lake Superior near Superior, Wisconsin. Although the majority of the spilled material evaporated, damage to aquatic life was extensive. Several thousand fishes were killed and an inestimable number were exposed to low concentrations (< 5 mg/L) of the chemical concentrate for several weeks. Fishes that survived the spill were examined within 7 days of exposure to determine the extent of injury when compared to fishes collected from the reference site. The liver, spleen, gill, and head kidney were examined for histopathology. Blood was collected to determine the severity of liver damage reflected by the presence of the serum enzymes (aspartate aminotransferase, alanine aminotransferase, and {delta}-glutamyl transferase). 14 refs., 1 fig., 1 tab.

  10. Exertional myopathy in whooping cranes (Grus americana) with prognostic guidlelines

    USGS Publications Warehouse

    Hanley, C.S.; Thomas, N.J.; Paul-Murphy, P.; Hartup, B.K.

    2005-01-01

    Exertional myopathy developed in three whooping cranes (Grus americana) secondary to routine capture, handling, and trauma. Presumptive diagnosis of exertional myopathy was based on history of recent capture or trauma, clinical signs, and elevation of aspartate aminotransferase, alanine aminotransferase, creatine kinase, lactate dehydrogenase, and serum potassium. Treatments were attempted in each case, but ultimately were not successful. Gross and microscopic lesions at necropsy confirmed the diagnosis in each case, with the leg musculature most severely affected. Guidelines for determining prognosis of exertional myopathy in cranes have been included based on the analysis of these cases and others in the literature. As treatment is largely unrewarding, prevention remains the key in controlling exertional myopathy. Identification of predisposing factors and proper handling, immobilization, and transportation techniques can help prevent development of exertional myopathy in cranes.

  11. The effects of ultraviolet C radiation on the ultrastructure of the liver cells of mole rats.

    PubMed

    Tekın, Saban; Türker, Hüseyin; Güven, Turan; Yel, Mustafa

    2016-01-01

    The aim of this study was to elucidate the ultrastructural changes in the liver cells of mole rats (Spalax leucodon) exposed to ultraviolet radiation (UVR). Thirteen mole rats used in this study were caught from nature. They were divided into four groups. The first group was separated as a control and was not given any radiation. The rest were exposed to ultraviolet C (UVC) radiation for 7, 14, and 21 days. The electron microscopic examinations revealed that significant ultrastructural changes occurred in the liver tissue. These changes were the reduction in cytoplasmic organelles, dilatation in rough endoplasmic reticulum, impairment of nucleus membrane, and broadened and vacuolated mitochondria in the cytoplasm. Also, UVC radiation caused significant changes in liver enzymes of aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and gama-glutamiltransferase values. After long-term exposure to radiation, some excessive ultrastructural changes occurred. These results indicated that longer exposure to UVR would cause more ultrastructural effects on the liver cells and liver enzymes.

  12. Experimental acute toxicity of xylitol in dogs.

    PubMed

    Xia, Z; He, Y; Yu, J

    2009-10-01

    The Cases of xylitol poisoning in dogs are increasing as a result of ingestion of xylitol-containing products. Eighteen adult, clinically normal Pekingese dogs were orally dosed with 1 or 4 g/kg xylitol in aqueous solution. Blood samples were collected before and after dosing. Plasma insulin concentrations of both treated groups rose sharply from 20 min after xylitol dosing, peaking at 40 min. Hypoglycemia followed the increase in insulin concentration, with blood glucose values started to decrease 30 min after dosing. Other plasma biochemistry changes associated with xylitol administration were increased alanine aminotransferase and aspartate aminotransferase activities, hypophosphatemia, hypokalemia, and hypercalcemia. Plasma sodium and chloride concentrations remained normal. This study established a biochemical basis for diagnosis and treatment of xylitol poisoning in dogs.

  13. Protective effect of alcoholic extract of Entada pursaetha DC. against CCl4-induced hepatotoxicity in rats.

    PubMed

    Gupta, Gaurav; More, Amar Sunil; Kumari, Rashmi Rekha; Lingaraju, Madhu Cholenahalli; Kumar, Dhirendra; Kumar, Dinesh; Mishra, Santosh Kumar; Tandan, Surender Kumar

    2014-03-01

    The alcoholic extract of stem of E. pursaetha (PSE, 30, 100, 300 mg/kg body weight, po for 7 days) showed hepatoprotective activity against CCl4 (2 mL/kg body weight, ip)-induced hepatotoxicity. The extract exhibited a significant dose-dependent hepatoprotective effect comparable to standard drug silymarin, by preventing increase in serum levels of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, total protein, and total bilirubin, lactate dehydrogenase; by lowering hepatic levels of malonaldehyde, nitrate-nitrite, myeloperoxidase activity; enhancing activities of antioxidant enzymes, superoxide dismutase, catalase and increasing reduced glutathione levels in liver, which suggests the antioxidant property of PSE. Histopathological studies also supported the above biochemical parameters. The results suggested that alcoholic extract of E. pursaetha possesses significant hepatoprotective activity in CCl4-induced acute hepatotoxicity in rats and this is likely to be mediated through its antioxidant activities.

  14. A blood chemistry profile for lake trout

    USGS Publications Warehouse

    Edsall, Carol Cotant

    1999-01-01

    A blood chemistry profile for lake trout Salvelinus namaycush was developed by establishing baseline ranges for several clinical chemistry tests (glucose, total protein, amylase, alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, creatine kinase, calcium, and magnesium). Measurements were made accurately and rapidly with a Kodak Ektachem DT60 Analyzer and the Ektachem DTSC Module. Blood serum was collected from both laboratory-reared lake trout (1978 and 1986 year-classes) and feral spawning trout from Lake Michigan and then analyzed in the laboratory. No clinically significant differences were found between samples analyzed fresh and those frozen for 1 or 6 weeks. The ranges in chemistry variables for feral lake trout were generally wider than those for laboratory-reared lake trout, and significant differences existed between male and female feral lake trout for several tests. Blood chemistry profiles also varied seasonally on fish sampled repeatedly.

  15. Machine perfusion versus cold storage of livers: a meta-analysis.

    PubMed

    Liu, Sushun; Pang, Qing; Zhang, Jingyao; Zhai, Mimi; Liu, Sinan; Liu, Chang

    2016-12-01

    Different organ preservation methods are key factors influencing the results of liver transplantation. In this study, the outcomes of experimental models receiving donation after cardiac death (DCD) livers preserved through machine perfusion (MP) or static cold storage (CS) were compared by conducting a meta-analysis. Standardized mean difference (SMD) and 95% confidence interval (CI) were calculated to compare pooled data from two animal species. Twenty-four studies involving MP preservation were included in the meta-analysis. Compared with CS preservation, MP can reduce the levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), and hyaluronic acid (HA) and the changes in liver weight. By contrast, MP can enhance bile production and portal vein flow (PVF). Alkaline phosphatase (ALP) levels and histological changes significantly differed between the two preservation methods. In conclusion, MP of DCD livers is superior to CS in experimental animals.

  16. Thermal deterioration of oil and frying foodstuffs.

    PubMed

    Totani, Nagao; Ojiri, Yuko

    2007-01-01

    Male Wistar rats were fed ad libitum for 12 weeks a powdered diet (AIN93G; no fat) containing 7 wt% of fresh oil (control), and frying oils heated for 20 h at 180 degrees C with amino acids, gluten, sugar, and wheat starch, respectively. The rats were subjected to anthropometric measurements, hematological analyses, and observations of the liver and kidneys. All of the rats grew well, and no gross symptoms attributable to the experimental oils were observed. The serum of all the experimental groups showed a tendency toward lower levels of triacylglycerol and free fatty acids and higher levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) than that of the control group. Among experimental groups, the rats fed a diet containing oil heated with gluten were least influenced by thermally deteriorated oil in terms of serum levels of glucose, triacylglycerol, phospholipids, cholesterol, and insulin; histological evaluations; and number of dark-red patches found on the surface of the liver.

  17. [Unexplained, subclinical chronically elevated transaminases].

    PubMed

    Vital Durand, D; Lega, J-C; Fassier, T; Zenone, T; Durieu, I

    2013-08-01

    Unexplained, subclinical chronically elevated transaminases is mainly a marker of non-alcoholic fatty liver disease, metabolic syndrome, alcoholism and diabetes, which are very common situations but viral hepatitis and iatrogenic origin must also be considered. Before looking for hepatic or genetic rare diseases, it is worth considering hypertransaminasemia as a clue for muscular disease, particularly in paediatric settings, and creatine phosphokinase is a specific marker. Then, patient history, examination and appropriate biologic requests can permit the identification of less frequent disorders where isolated hypertransaminasemia is possibly the unique marker of the disease for a long while: hemochromatosis, celiac disease, autoimmune hepatitis, Wilson's disease, α1-anti-trypsine deficiency, thyroid dysfunctions, Addison's disease. Liver biopsy should be performed only in patients with aspartate aminotransferases upper the normal range or alanine aminotransferases higher than twice the normal range after 6 months delay with dietetic corrections.

  18. Hepatoprotective activity of Moringa oleifera on antitubercular drug-induced liver damage in rats.

    PubMed

    Pari, L; Kumar, N Ashok

    2002-01-01

    Moringa oleifera Lam (Moringaceae), commonly known as "Drumstick," is used in Indian folk medicine for the treatment of various illness. We have evaluated the hepatoprotective effect of an ethanolic extract of M. oleifera leaves on liver damage induced by antitubercular drugs such as isoniazid (INH), rifampicin (RMP), and pyrazinamide (PZA) in rats. Oral administration of the extract showed a significant protective action made evident by its effect on the levels of glutamic oxaloacetic transaminase (aspartate aminotransferase), glutamic pyruvic transaminase (alanine aminotransferase), alkaline phosphatase, and bilirubin in the serum; lipids, and lipid peroxidation levels in liver. This observation was supplemented by histopathological examination of liver sections. The results of this study showed that treatment with M. oleifera extracts or silymarin (as a reference) appears to enhance the recovery from hepatic damage induced by antitubercular drugs.

  19. Liver injury suppressing compounds from avocado (Persea americana).

    PubMed

    Kawagishi, H; Fukumoto, Y; Hatakeyama, M; He, P; Arimoto, H; Matsuzawa, T; Arimoto, Y; Suganuma, H; Inakuma, T; Sugiyama, K

    2001-05-01

    To evaluate the protective activity of fruits against liver injury, 22 different fruits were fed to rats with liver damage caused by D-galactosamine, a powerful liver toxin. As measured by changes in the levels of plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST), avocado showed extraordinarily potent liver injury suppressing activity. Five active compounds were isolated and their structures determined. These were all fatty acid derivatives, of which three, namely, (2E,5E,12Z,15Z)-1-hydroxyheneicosa-2,5,12,15-tetraen-4-one, (2E,12Z,15Z)-1-hydroxyheneicosa-2,12,15-trien-4-one, and (5E,12Z)-2-hydroxy-4-oxoheneicosa-5,12-dien-1-yl acetate, were novel.

  20. Plasma biochemistry in the horse during 3-day event competition.

    PubMed

    Rose, R J; Ilkiw, J E; Arnold, K S; Backhouse, J W; Sampson, D

    1980-07-01

    Blood samples were collected from 16 Thoroughbred horses before, during and after the second day of a 3-day event. Plasma osmolality, concentrations of sodium, potassium, chloride, urea, creatinine, glucose, bilirubin, iron, total protein, albumin, alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, creatine kinase, calcium, inorganic phosphate, uric acid, cholesterol, triglycerides and non-esterified fatty acids were measured. Significant differences from pre-event values were found in all parameters with the greatest changes being found after the cross-country phase. Most parameters showed significant rises following exercise, except calcium and chloride, which decreased. It was deduced from the changes in biochemistry that dehydration, reduced glomerular filtration rate, increased glycogenolysis and increased lipid metabolism, were a result of this form of competitive exercise.

  1. Blood changes during training and racing in sled dogs.

    PubMed

    Querengaesser, A; Iben, C; Leibetseder, J

    1994-12-01

    Six male and six female Alaskan Huskies allocated three by three to two teams fed rations slightly different in protein/fat ratio (A, 29.2: 53.7%; B, 34.1: 48.5% of digestible energy) were studied for hematological and metabolic changes during a complete training and racing season (24 wk). Blood variables [packed cell volume (PCV), red blood cell count (RBC), mean corpuscular volume, hemoglobin, total plasma protein, free fatty acids, cholesterol, lactic acid, creatine phosphokinase (CPK), creatinine, alanine aminotransferase (ALT) and aspartate aminotransferase (AST)] and six minerals were determined before (at rest) and immediately after a test run of 9 km and then 10, 20 and 30 min later, at the beginning of training as well as 7, 20 and 24 wk later. Training and exercise both significantly influenced PCV, RBC, creatinine and ALT and exercise influenced CPK. Only negligible differences were found between the diets.

  2. Liver Protective Effects of Morinda citrifolia (Noni)

    PubMed Central

    Nowicki, Diane; Anderson, Gary; Jensen, Jarakae; West, Brett

    2008-01-01

    This study evaluated the protective effects of Noni fruit juice on acute liver injury induced by carbon tetrachloride (CCl4) in female Sprague-Dawley (SD) rats. Liver damage (micro-centrilobular necrosis) was observed in animals pretreated with 20% placebo (drinking water) + CCl4. However, pretreatment with 20% Noni juice in drinking water + CCl4 resulted in markedly decreased hepatotoxic lesions. Furthermore, serum alanine aminotransferase and aspartate aminotransferase levels were significantly lower in the Noni group than the placebo group. In a correlative time-dependent study, one dose of CCl4 (0.25 mL/kg in corn oil, p.o.) in female SD rats, pretreated with 10% placebo for 12 days, caused sequential progressive hepatotoxic lesions over a 24 h period, while a protective effect from 10% Noni juice pretreatment was observed. These results suggest that Noni juice is effective in protecting the liver from extrinsic toxin exposure. PMID:18317933

  3. Hematology and clinical chemistry values of normal and euthymic hairless adult male Dunkin-Hartley guinea pigs (Cavia porcellus).

    PubMed

    Waner, Trevor; Avidar, Yaakov; Peh, Hao-Chang; Zass, Rosa; Bogin, Eitan

    1996-01-01

    Hematology and serum chemistry measurements were performed on blood specimens from 12 male Dunkin-Hartley hairless guinea pigs Crl:IAF(HA)BR and 10 haired Dunkin-Hartley male guinea pigs Crl:(HA)BR. Significantly higher activities of alanine aminotransferase, aspartate aminotransferase, amylase, and creatine kinase were observed in the hairless guinea pigs as compared to the haired strain. Alkaline phosphatase activity was found to be lower in the hairless guinea pig. The hairless guinea pigs were found to have serum urea concentrations approximately 46% higher than the normal guinea pig strain. The erythrocytic mean cell volume of the hairless strain was found to be smaller, with a greater hemoglobin content. Hairless guinea pigs were found to have approximately 40% fewer leukocytes with a reversed lymphocyte:neutrophil ratio compared to the haired guinea pigs which had much higher lymphocyte counts.

  4. Plasma cholesterol and triglycerides in heroin addicts.

    PubMed

    Maccari, S; Bassi, C; Zanoni, P; Plancher, A C

    1991-12-31

    We examined total cholesterolemia, triglyceridemia, high density lipoproteins- (HDL) cholesterolemia, apolipoproteins A1 and B, body mass index, albuminemia and alanine aminotransferase in 60 heroin addicts. After comparing 23 control subjects with the heroin addicts the result was that the latter have significantly lower mean values of total cholesterolemia and of HDL-cholesterolemia and higher values of triglyceridemia. They also have significantly higher prevalences of cases of hypocholesterolemia and of hypo-HDL-cholesterolemia. Within the addict group there is no linear correlation between total cholesterolemia and body mass index; there is, however, an inverse linear correlation between total cholesterolemia and alanine aminotransferase. Therefore, the alterations found in the lipid pattern of heroin addicts are not due to malnutrition but hypothetically to liver diseases which are frequent in these patients.

  5. Clinical Chemistry of Patients With Ebola in Monrovia, Liberia.

    PubMed

    de Wit, Emmie; Kramer, Shelby; Prescott, Joseph; Rosenke, Kyle; Falzarano, Darryl; Marzi, Andrea; Fischer, Robert J; Safronetz, David; Hoenen, Thomas; Groseth, Allison; van Doremalen, Neeltje; Bushmaker, Trenton; McNally, Kristin L; Feldmann, Friederike; Williamson, Brandi N; Best, Sonja M; Ebihara, Hideki; Damiani, Igor A C; Adamson, Brett; Zoon, Kathryn C; Nyenswah, Tolbert G; Bolay, Fatorma K; Massaquoi, Moses; Sprecher, Armand; Feldmann, Heinz; Munster, Vincent J

    2016-10-15

    The development of point-of-care clinical chemistry analyzers has enabled the implementation of these ancillary tests in field laboratories in resource-limited outbreak areas. The Eternal Love Winning Africa (ELWA) outbreak diagnostic laboratory, established in Monrovia, Liberia, to provide Ebola virus and Plasmodium spp. diagnostics during the Ebola epidemic, implemented clinical chemistry analyzers in December 2014. Clinical chemistry testing was performed for 68 patients in triage, including 12 patients infected with Ebola virus and 18 infected with Plasmodium spp. The main distinguishing feature in clinical chemistry of Ebola virus-infected patients was the elevation in alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and γ-glutamyltransferase levels and the decrease in calcium. The implementation of clinical chemistry is probably most helpful when the medical supportive care implemented at the Ebola treatment unit allows for correction of biochemistry derangements and on-site clinical chemistry analyzers can be used to monitor electrolyte balance.

  6. Protective effects of C-phycocyanin on alcohol-induced acute liver injury in mice

    NASA Astrophysics Data System (ADS)

    Xia, Dong; Liu, Bing; Luan, Xiying; Sun, Junyan; Liu, Nana; Qin, Song; Du, Zhenning

    2016-03-01

    Excessive alcohol consumption leads to liver disease. Extensive evidence suggests that C-phycocyanin (C-PC), a chromophore phycocyanobilin derived from Spirulina platensis, exerts protective effects against chemical-induced organ damage. In this study, we investigated whether C-PC could protect against ethanol-induced acute liver injury. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (CHOL), low-density lipoprotein (LDL), liver homogenate malondialdehyde (MDA), superoxide dismutase (SOD) content were measured, and pathological examination of liver sections were examined. C-PC showed obvious inhibitory effects on serum ALT, AST, TG, CHOL, LDL and MDA, and SOD content significantly increased in the liver. The structure of hepatic lobules was clear, liver sinus returned to normal, and liver cell cords were arranged in neat rows. Cloudiness, swelling, inflammatory cell infiltration and spotty necrosis of liver cells were significantly reduced. Therefore, C-PC can significantly protect against ethanol-induced acute liver injury.

  7. Intestinal Hypoganglionosis Leading to Intestinal Failure and the Compassionate Use of Omegaven™

    PubMed Central

    Karjoo, Sara; Danielson, Paul; Wilsey, Michael; Shakeel, Fauzia

    2017-01-01

    Intestinal hypoganglionosis is a rare innervation disorder that provides numerous nutritional, medical and surgical challenges. In this case report, we present a case of a newborn with intestinal hypoganglionosis leading to intestinal failure and intestinal failure-associated liver disease who responded to Omegaven™, a fat emulsion comprised of omega-3 fatty acids. Omegaven™ has been shown to be beneficial in the management of cholestatic liver injury. Clinical success with Omegaven™ was seen in this patient with a clear decrease in aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase and complete resolution of cholestasis with a direct bilirubin of zero within two weeks of initiation of Omegaven™. No current guidelines for the diagnosis and management of hypoganglionosis are available. We recommend a multidisciplinary approach and the use of novel therapies such as fat emulsions composed of omega-3 fatty acids for improved patient outcomes. Appropriate compassionate use protocols should be obtained from the Food and Drug Administration prior to initiation of Omegaven™.

  8. Serum biochemistry in Andean flamingos (Phoenicoparrus andinus): natural versus artificial diet.

    PubMed

    Norambuena, M Cecilia; Parada, Mario

    2005-09-01

    A study of 10 clinical pathology values in four groups of Andean flamingo chicks (Phoenicoparrus andinus) was conducted to evaluate an artificial feeding program in Chile. Three groups were fed controlled diets (groups 2000, 2001, and 2002) with quantitative differences in their nutritional content. A fourth group of free-living Andean flamingo chicks was used as normal controls. Nutritional management techniques used in 2002 resulted in hematologic values with similar levels of total protein, globulins, albumin, cholesterol, urea, phosphorus, aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatinine, and calcium to those obtained in free-living chicks. In addition, final weight, physical condition, and plumage in flamingo chicks of group 2002 were considered satisfactory to face local climatic conditions and nomadic activity. These results may be useful as reference values and help to improve conservation management and veterinary care of this species.

  9. Subcellular Distribution of Enzymes of Glycolate Metabolism in the Alga Cyanidium caldarium1

    PubMed Central

    Gross, Wolfgang; Beevers, Harry

    1989-01-01

    The intracellular distribution of enzymes capable of catalyzing the reactions from phosphoglycolate to glycerate in the bluegreen colored eucaryotic alga Cyanidium caldarium has been studied. After separating the organelles from a crude homogenate on a linear flotation gradient, the enzymes glycolate oxidase and glutamate-glyoxylate aminotransferase along with catalase were present in the peroxisomal fraction (density: 1.23 grams per cubic centimeter). Serine hydroxymethyltransferase was found in the mitochondrial fraction (density: 1.18 grams per cubic centimeter). In contrast to the observations in green leaves of higher plants, the enzymes for the conversion of serine to glycerate (serine-glyoxylate aminotransferase and hydroxypyruvate reductase) were found only in the soluble fraction of the gradient. The partial characterization of enzymes from Cyanidium participating in glycolate metabolism revealed only slight differences from the corresponding enzymes from higher plants. The phylogenetic implications of the observed similarities between the enigmatic alga Cyanidium and higher plants are discussed. PMID:16666880

  10. Enzymatic activities in brains of diabetic rats treated with vanadyl sulphate and sodium tungstate.

    PubMed

    Lemberg, A; Fernández, M A; Ouviña, G; Rodríguez, R R; Peredo, H A; Susemihl, C; Villarreal, I; Filinger, E J

    2007-12-01

    The hypothesis of the present study was that diabetes mellitus might affect brain metabolism. Streptozotocin (STZ)-induced diabetic rats, treated with vanadyl sulphate (V) and sodium tungstate (T) were employed to observe the aspartate aminotransferase (AST), alanine aminotransferase (ALT) and creatine kinase (CK) activities in brain homogenates. Significant increases in AST, ALT and CK activities were found in diabetic brain homogenates against controls, suggesting increments of transamination in brain and/or increases in cell membrane permeability to these enzymes. The increase in brain CK possibly expresses alterations in energy production. The decrease in CK activity caused by V and T treatment in diabetic rats suggests that both agents tend to normalize energy consumption. It is also possible that V and T-induced hypoglycemic effects cause metabolic alterations in brain.

  11. Variation in metabolic enzymatic activity in white muscle and liver of blue tilapia, Oreochromis aureus, in response to long-term thermal acclimatization

    NASA Astrophysics Data System (ADS)

    Younis, Elsayed M.

    2015-05-01

    The effects of rearing temperature on white muscle and hepatic phosphofructokinase (PFK), pyruvate kinase (PK), lactate dehydrogenase (LDH), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were examined in fingerlings of blue tilapia, Oreochromis aureus. The experiment was conducted for 14 weeks at temperatures of 18, 22, 26, 30, and 34°C. The activity of the glycolytic enzymes PFK, PK, and LDH in white muscle increased significantly with increase in water temperature. A reverse trend was observed for these enzymes in the liver, except for LDH, which behaved in the same manner as in white muscle. Cytosolic AST and ALT activity increased in both white muscle and liver in response to warm thermal acclimatization, while a reduction in mitochondrial AST and ALT activity was noticed at high temperatures in comparison with those at a lower temperature.

  12. Enzyme activities in plasma, kidney, liver, and muscle of five avian species

    USGS Publications Warehouse

    Franson, J.C.; Murray, H.C.; Bunck, C.

    1985-01-01

    Activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), creatine phosphokinase (CPK), and lactate dehydrogenase (LDH) were determined in plasma, kidney, liver, and muscle from five species of captive birds. Few differences occurred in plasma activities between sexes but considerable differences occurred between species. All five enzymes were detected in each of the tissues sampled. Relative enzyme activities in liver, kidney, and muscle were similar for each species. CPK activity was much higher in muscle than in liver or kidney and, of the five enzymes studied, may be the best indicator of muscle damage. Most of the other enzymes were more evenly distributed among the three tissues, and no organ-specific enzyme could be identified for liver or kidney. Because of interspecific variations in plasma enzyme activities, it is important to establish baseline values for each species to ensure accurate interpretation of results.

  13. Liver protective effects of Morinda citrifolia (Noni).

    PubMed

    Wang, Mian-Ying; Nowicki, Diane; Anderson, Gary; Jensen, Jarakae; West, Brett

    2008-06-01

    This study evaluated the protective effects of Noni fruit juice on acute liver injury induced by carbon tetrachloride (CCl(4)) in female Sprague-Dawley (SD) rats. Liver damage (micro-centrilobular necrosis) was observed in animals pretreated with 20% placebo (drinking water) + CCl(4). However, pretreatment with 20% Noni juice in drinking water + CCl(4) resulted in markedly decreased hepatotoxic lesions. Furthermore, serum alanine aminotransferase and aspartate aminotransferase levels were significantly lower in the Noni group than the placebo group. In a correlative time-dependent study, one dose of CCl(4) (0.25 mL/kg in corn oil, p.o.) in female SD rats, pretreated with 10% placebo for 12 days, caused sequential progressive hepatotoxic lesions over a 24 h period, while a protective effect from 10% Noni juice pretreatment was observed. These results suggest that Noni juice is effective in protecting the liver from extrinsic toxin exposure.

  14. Altered muscle energy metabolism in post-absorptive patients with chronic renal failure.

    PubMed

    Pastoris, O; Aquilani, R; Foppa, P; Bovio, G; Segagni, S; Baiardi, P; Catapano, M; Maccario, M; Salvadeo, A; Dossena, M

    1997-06-01

    Skeletal muscle biopsies were performed on 12 healthy sedentary subjects and on 22 non-dyalized chronic renal failure patients (CRF) on a free diet and after overnight fasting. Parathormone, glucagon and insulin were determined at the same time of biopsies. CRF patients showed significantly low ATP and creatine phosphate levels. Regarding enzyme activities, a high hexokinase Vmax was found, while the pyruvate kinase activity was lower than in the control group. For the tricarboxylic acid cycle, citrate synthase, succinate dehydrogenase and malate dehydrogenase activities were higher; total NADH cytochrome c reductase activity was also high, while cytochrome oxidase activity was slightly lower. Both alanine aminotransferase and aspartate aminotransferase activities were considerably high in comparison with the control group. In conclusion, our study revealed a hypermetabolic TCA cycle, but impaired oxidative phosphorylation, which partly explained the reduced ATP concentration. Excessive protein intake and hormonal derangements may play a role in these metabolic changes.

  15. An In Vitro and In Vivo Study of the α-Amylase Activity of Phaseolamin

    PubMed Central

    de Gouveia, Neire Moura; Alves, Fernanda Vieira; Furtado, Fabiana Barcelos; Scherer, Danielli Luana; Mundim, Antonio Vicente

    2014-01-01

    Abstract We evaluated the polypeptide profiles, inhibition of human salivary α-amylase activity, and hemagglutination properties of a commercial phaseolamin sample. We also performed an in vivo assay to investigate the effects of a commercial phaseolamin treatment (100, 500, or 1500 mg/kg) over 20 days on the glycemia, body weight, and serum biochemical parameters (total cholesterol, triglycerides, alanine aminotransferase, and aspartate aminotransferase) of nondiabetic and streptozotocin-induced diabetic rats. The in vitro evaluation showed defined protein profiles, low hemagglutination activity, and high α-amylase inhibition. None of the experimental groups treated with phaseolamin or acarbose showed decreases in body weight. Our data demonstrate that phaseolamin inhibits amylase activity in vitro, reduces blood glucose levels, decreases or attenuates some of the renal and hepatic effects of diabetes in streptozotocin-induced rats, and could therefore have therapeutic potential in the treatment or prevention of the complications of diabetes. PMID:24650210

  16. Tolerance of adult mallards to subacute ingestion of crude petroleum oil

    USGS Publications Warehouse

    Rattner, B.A.

    1981-01-01

    Adult male mallards were fed untreated mash or mash containing 1.5% Prudhoe Bay crude oil for 7 days ad lib. During the initial 24 h of exposure to crude petroleum oil, ducks consumed less mash (P less than 0.05) and lost approx. 3.5% of their initial body weight (P less than 0.05), however, neither intake nor body weight differ between groups on days 2-7. Plasma samples collected between 09.00 and 10.00 h on days 0, 1, 3, or 7 indicated that corticosterone, glucose, thyroxine, total protein, and uric acid concentrations, and the activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and butyrylcholinesterase (BCHE) were not affected by treatment. These findings suggest that adult mallards may be able to tolerate large quantities of crude petroleum oil mixed in their diet (approx. 25 ml over a 7-day period) without overt or biochemical indications of distress.

  17. IFCC reference procedures for measurement of the catalytic concentrations of enzymes: corrigendum, notes and useful advice. International Federation of Clinical Chemistry and Laboratory Medicine (IFCC)--IFCC Scientific Division.

    PubMed

    Schumann, Gerhard; Canalias, Francesca; Joergensen, Poul J; Kang, Dongchon; Lessinger, Jean-Marc; Klauke, Rainer; Committee On Reference Systems For Enzymes C-Rse; International Federation of Clinical Chemistry and Laboratory Medicine Scientific Division

    2010-05-01

    The primary reference measurement procedures (PRMPs) for the international standardization of catalytic concentration measurements of alpha-amylase, alanine aminotransferase, aspartate aminotransferase (AST), creatine kinase (CK), gamma-glutamyltransferase and lactate dehydrogenase have been performed in reference laboratories for several years. The IFCC Committee on Reference Systems for Enzymes and two reference laboratories, with official accreditation for the PRMPs, have collected useful information on some of the steps of the reference procedures that require special attention. This document comprises errata corrige for minor mistakes in published PRMPs for AST and CK. Several notes on the PRMPs are emphasized. This includes details that are very important for improved standardization, and general suggestions for reducing measurement uncertainty.

  18. Aromatic hydrocarbon pathology in fish following a large spill into the Nemadji River, Wisconsin, USA

    USGS Publications Warehouse

    Caldwell, C.A.

    1997-01-01

    On June 30, 1992, a train accident resulted in a rail car releasing 114,000 L of a complex mixture of aromatic hydrocarbons into the Nemadji River, a tributary of Lake Superior near Superior, Wisconsin (Table 1). Although the majority of the spilled material evaporated, damage to aquatic life was extensive. Several thousand fishes were killed and an inestimable number were exposed to low concentrations (< 5 mg/L) of the chemical concentrate for several weeks (Allen 1993). Fishes that survived the spill were examined within 7 days of exposure to determine the extent of injury when compared to fishes collected from the reference site. The liver, spleen, gill, and head kidney were examined for histopathology. Blood was collected to determine the severity of liver damage reflected by the presence of the serum enzymes (aspartate aminotransferase, alanine aminotransferase, and d - glutamyl transferase).

  19. Antioxidant and hepatoprotective effects of punicalagin and punicalin on acetaminophen-induced liver damage in rats.

    PubMed

    Lin, C C; Hsu, Y F; Lin, T C; Hsu, H Y

    2001-05-01

    Punicalagin and punicalin were isolated from the leaves of Terminalia catappa L., a Combretaceous plant distributed throughout tropical and subtropical beaches, which is used for the treatment of dermatitis and hepatitis. Our previous studies showed that both of these compounds exert antioxidative activity. In this study, the antihepatotoxic activity of punicalagin and punicalin on acetaminophen-induced toxicity in the rat liver was evaluated. After evaluating the changes of several biochemical functions in serum, the levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were increased by acetaminophen administration and reduced by punicalagin and punicalin. Histological changes around the hepatic central vein and oxidative damage induced by acetaminophen were also recovered by both compounds. The data show that both punicalagin and punicalin exert antihepatotoxic activity, but treatment with larger doses enhanced liver damage. These results suggest that even if punicalagin and punicalin have antioxidant activity at small doses, treatment with larger doses will possibly induce some cell toxicities.

  20. [The effect of chlorpromazine and carbamazepine on diagnostically relevant liver enzymes].

    PubMed

    Göckeritz, W; Borchert, H H

    1990-07-01

    No interactions related to the analytical method were observed between chlorpromazine (1) or carbamazepine (2) and activities of alanine aminotransferase (ALAT), aspartate aminotransferase (ASAT), glutamate dehydrogenase (GLDH) or lactate dehydrogenase (LDH). With respect to its cytotoxic potential 1 in cultures of isolated rat hepatocytes increased markedly the release of enzymes into the culture medium, whereas the overall activities of the enzymes were not influenced. 2 in cultured hepatocytes caused no significant effects on the activities of the enzymes investigated. Besides the investigation of methodically related interactions in pooled human serum the methodic procedure including the use of cultures of isolated hepatocytes allows to study also pharmacologically and toxicologically related interactions between drugs and diagnostically relevant liver enzymes.

  1. Atherosclerosis and Liver Function Tests in Coronary Angiography Patients

    PubMed Central

    Doganer, YC; Rohrer, JE; Aydogan, U; Agerter, DC; Cayci, T; Barcin, C

    2015-01-01

    ABSTRACT Objective: Elevated aminotransferase levels indicating liver function, even in the normal range, have attracted great concern as potential novel markers of cardiovascular risk assessment. We hypothesized the possibility that liver function test variations in the normal range might be meaningfully associated to coronary artery disease (CAD). Method: Eighty-eight patients were randomly selected from those who underwent coronary angiography from June 2010 to June 2011 after applying to the outpatient cardiology clinic in Gulhane Military Medical Academy. According to the results of angiographies, patients were classified into three groups as normal, non-critical (< 50% involvement in coronaries), and critical (≥ 50% involvement in coronaries). In addition to angiographic intervention, measurements of serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) concentrations, albumin and the other serum parameters were performed in all patients. Results: The patient groups of CAD were balanced (28 critical cases, 30 non-critical cases and 30 normal cases). Mean age was 51.93 ± 9.3 (range 32–65) years and 19.3 per cent (n = 17) were females. Multiple linear regression analysis of all three liver function tests explained a significant portion of the variance, but adjusted r-squares were small (AST = 0.174, ALT = 0.242, albumin = 0.124). Albumin was significantly higher for patients with critical CAD than for patients with no CAD (beta = 3.205, p = 0.002). Non-critical CAD was not significantly different from no CAD for any of the dependent variables. Mean AST was significantly higher for patients taking aspirin (beta = 0.218, p = 0.049), as was mean ALT (beta = 0.264, p = 0.015). Conclusion: Alanine aminotransferase and AST may not be associated with angiographically determined coronary atherosclerosis. Albumin may be more sensitive to demonstrate the burden of atherosclerosis. These results indicate that the association between the liver

  2. Hepatoprotective Effects of Corilagin Following Hemorrhagic Shock are Through Akt-Dependent Pathway

    PubMed Central

    Liu, Fu-Chao; Chaudry, Irshad H.; Yu, Huang-Ping

    2017-01-01

    ABSTRACT Corilagin, a component of Phyllanthus urinaria extract, possesses antioxidant, thrombolytic, antiatherogenic, and hepatoprotective properties, but the mechanism underlying these effects remains unclear. Previous studies showed that the Akt (protein kinase B) signaling pathway exerts anti-inflammatory and organ protective effects. The aim of this study was to investigate the mechanism of action of corilagin and determine whether these effects are mediated through the Akt-dependent pathway in a trauma-hemorrhagic shock-induced liver injury rodent model. Hemorrhagic shock was induced in male Sprague–Dawley rats; mean blood pressure was maintained at 35 mm Hg to 40 mm Hg for 90 min, followed by fluid resuscitation. During resuscitation, three doses of corilagin alone (1 mg/kg, 5 mg/kg, or 10 mg/kg, intravenously) were administered. Furthermore, a single dose of corilagin (5 mg/kg) with and without Wortmannin (1 mg/kg, PI3K inhibitor), Wortmannin alone, or vehicle was administered. Twenty-four hours after resuscitation, plasma alanine aminotransferase and aspartate aminotransferase concentration and hepatic parameters were measured. One-way ANOVA was used for statistical analysis. Hepatic myeloperoxidase activity and the concentrations of plasma alanine aminotransferase and aspartate aminotransferase, interleukin-6, tumor necrosis factor-α, intercellular adhesion molecule-1, and cytokine-induced neutrophil chemoattractant-1 (CINC-1) and CINC-3 increased following hemorrhagic shock. These parameters were significantly attenuated in corilagin-treated rats following hemorrhagic shock. Hepatic phospho-Akt expression was also higher in corilagin-treated rats than in vehicle-treated rats. The elevation of phospho-Akt was abolished by combined treatment with Wortmannin and corilagin. Our results suggest that corilagin exerts its protective effects on hemorrhagic shock-induced liver injury, at least, via the Akt-dependent pathway. PMID:27559697

  3. Pharmacokinetic Modeling of Trivalent and Hexavalent Chromium Based on Ingestion and Inhalation of Soluble Chromium Compounds.

    DTIC Science & Technology

    1991-12-01

    located primarily in the epiphyseal region of -he long bones. The distribution of radiolabel was reported to be similar to that seen after administration...of bone-seeking tracers 9 like Ca45 or Sr 89. Kraintz and Talmage (1952) had also noted localization of radiolabel in the epiphyses of long bones...weight gain, histologic findings, red and white blood cell counts, serum activities of alanine aminotransferase, alkaline phosphatase, total cholesterol

  4. Changes of biochemical parameters and enzyme activities in broiler chickens with cold-induced ascites.

    PubMed

    Daneshyar, M; Kermanshahi, H; Golian, A

    2009-01-01

    An experiment with 250 one-day-old male broilers (Ross 308) was conducted to investigate the differences of some blood parameters of cold-induced ascitic and healthy broiler chicks in a 6-wk period. The chickens were divided into 2 groups of 5 replicates each. One group of these chickens was raised in normal temperature (NT) treatment and the other in cold temperature (CT) treatment to induce ascites. Mortality was necropsied daily to determine cause of death. At the end of the experiment (wk 6), 5 chickens from each replicate were randomly selected and slaughtered. The heart was removed; the right ventricle was dissected away from the left ventricle and septum. Weights of right and left ventricles were determined separately. Average BW gain and average feed intake were measured weekly, and weekly average feed conversion ratio was calculated. Serum glucose, total protein, cholesterol, triglyceride, activity of lactate dehydrogenase, as-partate aminotransferase, and alanine aminotransferase were determined. Throughout the study, the right ventricle-to-total ventricle ratio and total mortality percentage due to ascites of CT-treated birds at the end of experiment was greater (P < or = 0.05) than those of NT-treated ones. Fasting blood sugar of CT-treated birds in wk 4 and 6 was greater (P < or = 0.05) than NT-treated birds. Total blood protein of CT treatment was lower than NT-treated birds in every week and whole period, but this difference was only significant (P < or = 0.05) in wk 6. There was not a significant difference between 2 treatments for triglyceride and cholesterol, lactate dehydrogenase, aspartate aminotransferase, and alanine aminotransferase. It was concluded that cold-induced ascites could affect serum protein and fasting blood sugar of broiler chickens.

  5. Systemic and Pulmonary Hypertension After Resuscitation with Cell-Free Hemoglobin

    DTIC Science & Technology

    1992-07-01

    according to published standards (24). Inorganic phosphate was measured on a blood chemistry analyzer (Cobas-Fara; Roche Diagnostic Systems, Nutley, NJ...alanine aminotransferase (ALT), and lactate dehydrogenase (LDH) on the blood chemistry analyzer . The final 4 ml of blood was heparinized, centrifuged...milliliter of blood was placed in 2 ml of ice-cold perchioric acid (70% wt/vol), which was used to determine whole blood lactate in the blood chemistry

  6. Single or combined effects of mannan-oligosaccharides and probiotic supplements on the total oxidants, total antioxidants, enzymatic antioxidants, liver enzymes, and serum trace minerals in cyclic heat-stressed broilers.

    PubMed

    Sohail, M U; Rahman, Z U; Ijaz, A; Yousaf, M S; Ashraf, K; Yaqub, T; Zaneb, H; Anwar, H; Rehman, H

    2011-11-01

    This study was intended to explore the oxidative status of broilers under cyclic heat stress (HS) as modulated by supplementation of mannan-oligosaccharides (MOS) and a probiotic mixture (PM). Two hundred fifty 1-d-old chicks were randomly divided into 5 groups. From d 22, birds were either kept in a thermoneutral zone (TN; 26.7°C for 24 h/d) or subjected to HS (35 ± 1.1°C and 75 ± 5% RH for 8 h/d from 1000 to 1800 h) to the conclusion of the study on d 42. Birds were fed either a corn-based diet (TN and HS groups) or the same diet supplemented with 0.5% MOS (HS-MOS group), 0.1% PM (HS-PM group), or their combination as a symbiotic (SYN; HS-SYN group). On d 42, birds were killed by cervical dislocation to collect serum for the determination of total oxidants, total antioxidants, paraoxonase, arylesterase, ceruloplasmin, aspartate aminotransferase, alanine aminotransferase, and trace minerals. Heat stress increased (P < 0.05) total oxidants and total antioxidants and decreased (P < 0.05) paraoxonase and arylesterase, with no change in ceruloplasmin, aspartate aminotransferase, and alanine aminotransferase activities. Dietary supplementation decreased (P < 0.05) total oxidants and total antioxidants, with no effect on the activities of other enzymes. Heat stress did not influence serum copper, zinc, and manganese concentrations of birds when compared with those in the TN group. However, MOS increased (P < 0.05) concentrations of all the trace minerals, whereas SYN increased (P < 0.05) concentrations of only zinc and copper. It was concluded that MOS or PM supplementation, alone or as a SYN, may reduce some of the detrimental effects of HS, whereas MOS alone or as a SYN may improve the absorption of trace minerals.

  7. [Modest clinical benefits of ASAT analysis in today's health care. A case report of macro-ASAT giving rise to pathologically increased ASAT levels].

    PubMed

    Larsson, Anders; Karlsson, Bengt

    2002-09-19

    Aspartate aminotransferase (AST) may occur as a macroenzyme after forming a complex with immunoglobulins. The complex results in decreased plasma clearance and thus elevated AST levels. So far, macro-AST has only been described in a few patients. Here, we report a case of macro-AST in a healthy female. Clinicians should be aware of this phenomenon so that patients are not subjected to unnecessary investigations.

  8. Relationships between lifestyle patterns and cardio-renal-metabolic parameters in patients with type 2 diabetes mellitus: A cross-sectional study

    PubMed Central

    Ogihara, Takeshi; Osonoi, Yusuke; Osonoi, Takeshi; Saito, Miyoko; Tamasawa, Atsuko; Nakayama, Shiho; Someya, Yuki; Ishida, Hidenori; Gosho, Masahiko; Kanazawa, Akio; Watada, Hirotaka

    2017-01-01

    Introduction While individuals tend to show accumulation of certain lifestyle patterns, the effect of such patterns in real daily life on cardio-renal—metabolic parameters remains largely unknown. This study aimed to assess clustering of lifestyle patterns and investigate the relationships between such patterns and cardio-renal-metabolic parameters. Participants and methods The study participants were 726 Japanese type 2 diabetes mellitus (T2DM) outpatients free of history of cardiovascular diseases. The relationship between lifestyle patterns and cardio-renal-metabolic parameters was investigated by linear and logistic regression analyses. Results Factor analysis identified three lifestyle patterns. Subjects characterized by evening type, poor sleep quality and depressive status (type 1 pattern) had high levels of HbA1c, alanine aminotransferase and albuminuria. Subjects characterized by high consumption of food, alcohol and cigarettes (type 2 pattern) had high levels of γ-glutamyl transpeptidase, triglycerides, HDL-cholesterol, blood pressure, and brachial-ankle pulse wave velocity. Subjects characterized by high physical activity (type 3 pattern) had low uric acid and mild elevation of alanine aminotransferase and aspartate aminotransferase. In multivariate regression analysis adjusted by age, gender and BMI, type 1 pattern was associated with higher HbA1c levels, systolic BP and brachial-ankle pulse wave velocity. Type 2 pattern was associated with higher HDL-cholesterol levels, triglycerides, aspartate aminotransferase, ɤ- glutamyl transpeptidase levels, and diastolic BP. Conclusions The study identified three lifestyle patterns that were associated with distinct cardio-metabolic-renal parameters in T2DM patients. Trial registration UMIN000010932 PMID:28273173

  9. [Activity of the marker liver enzymes under the conditions of toxic hepatitis and alimentary deprivation of protein].

    PubMed

    Voloshchuk, O N; Kopyl'chuk, G P; Buchkovskaia, I M

    2014-01-01

    The activity of the sorbitoldehydrogenase (SDH), alanine aminotransferase (ALT) and alkaline phosphatase (ALP) in the blood serum of rats with acetaminophen-induced hepatitis under the conditions of alimentary deprivation of protein was studied. The animals were divided into 3 groups: 1--rats with acute acetaminophen-induced hepatitis, maintained on the full ration; 2--rats with acute acetaminophen-induced hepatitis, maintained under the conditions of alimentary deprivation of protein; 3--control. The activity of the sorbitol dehydrogenase in blood serum was determined by the kinetic method, activity of the alanine aminotransferase and alkaline phosphatase - photometrically. It is shown, that in animals with the model hepatitis the activity of sorbitol dehydrogenase in blood serum increases 20-fold, wherein statistical significance between animals with hepatitis maintained under the conditions of full ration and those of low-protein diet is not established. In the group of animals with acetaminophen-induced hepatitis the preservation on the control level of the alkaline phosphatase activity on the base of the increase of alanine aminotransferase by 2.2 times and ratio ALT/ALP>5 testifies about hepatocellular liver injury. In the group of animals with drug-induced hepatitis and alimentary deprivation of protein, the increase of the alkaline phosphatase and alanine aminotransferase activity is observed, herewith the ratio ALT/ALP ranges from 2 to 5 and testifies about mixed liver injury. The conclusion was made, that alimentary deprivation of protein is the critical factor for the development of the disturbances of functional and structural liver integrity, and the therapeutic approaches to the correction of the drug-induced liver injury should be different depending on the value of protein ration in the anamnesis, taking into account the different types of liver injury.

  10. Response of blood serum constituents to production of and recovery from a kwashiorkor-like syndrome in the young pig.

    PubMed

    Pond, W G; Ellis, K J; Schoknecht, P

    1992-09-01

    Twenty-six 3-week-old genetically obese pigs were fed in two experiments to determine the serum chemistry profile during severe protein malnutrition and repletion. Severe protein deficiency was produced in pigs fed the high-fat, low-protein diet (growth failure, rough hair, low serum total protein and albumin). In Experiment 1, blood was sampled from the anterior vena cava of each pig five times during depletion and three times during repletion to determine serum total cholesterol, high density lipoprotein (HDL)-cholesterol, triglycerides, total protein, albumin, glucose, Ca, inorganic P, Mg, Na, K, Cl, total bilirubin, urea N, creatinine, alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, and gamma-glutamyltransferase. In Experiment 2, blood was sampled weekly for 8 weeks for serum total cholesterol, HDL-cholesterol, triglycerides, albumin, glucose, Ca, P, Mg and alkaline phosphatase. HDL-cholesterol was increased (P less than 0.01) and albumin was decreased (P less than 0.01) in protein-deficient pigs in both experiments. Creatinine, total bilirubin, gamma-glutamyltransferase, alanine aminotransferase, and aspartate aminotransferase were elevated in protein-deficient pigs compared with controls after 7 weeks of depletion. Inorganic P (P less than 0.01), Ca (P less than 0.01), and Mg (P less than 0.05) concentrations were depressed in protein-depleted pigs compared with controls in both experiments. After 8 weeks of repletion in Experiment 1, all elements except inorganic P were similar in the two groups. Short-term, severe, protein malnutrition affected lipid, electrolyte, and structural mineral metabolism and indices of liver function in the absence of parasites, diarrhea, and infection. The effects were reversed after 8 weeks of repletion. We conclude that the elevated serum cholesterol in protein deficiency is related primarily to an increase in the HDL fraction.

  11. Acute hepatitis in a woman following excessive ingestion of an energy drink: a case report

    PubMed Central

    2011-01-01

    Introduction The consumption of energy drinks has increased significantly. We report the case of a patient who presented to our hospital with jaundice, abdominal pain, and markedly increased liver transaminases likely due to the increased consumption of an energy drink. To the best of our knowledge, this is the first case report in the literature linking the development of acute hepatitis to the consumption of an energy drink. Case presentation A 22-year-old Caucasian woman presented to our hospital with epigastric pain, nausea, vomiting, and low-grade fever. She had been drinking 10 cans of an energy drink daily for two weeks prior to presentation. Her physical examination revealed mild epigastric tenderness. Her initial blood tests revealed elevated alanine aminotransferase, aspartate aminotransferase, and total bilirubin. A computed tomographic scan of the abdomen and pelvis was normal, and the patient was discharged to home. She returned to the Emergency Department of our hospital with worsening pain and new-onset jaundice. This time her physical examination revealed epigastric tenderness and icteric sclera. Her aspartate aminotransferase, alanine aminotransferase, and international normalized ratio were markedly elevated. Further radiological studies were non-specific, and she was admitted to our hospital with a diagnosis of acute hepatitis. Her viral serology and toxicology screens were negative. The patient was treated supportively and was discharged after resolution of her symptoms and a marked decrease in her liver enzymes. Conclusion The development of acute hepatitis in this patient was most likely due to the excessive ingestion of an energy drink, and we speculate that niacin was the culprit ingredient. PMID:21696583

  12. Ongoing liver inflammation in patients with chronic hepatitis C and sustained virological response

    PubMed Central

    Welsch, Christoph; Efinger, Mira; von Wagner, Michael; Herrmann, Eva; Zeuzem, Stefan

    2017-01-01

    Background Novel direct-acting antiviral DAA combination therapies tremendously improved sustained virologic response (SVR) rates in patients with chronic HCV infection. SVR is typically accompanied by normalization of liver enzymes, however, hepatic inflammation, i.e. persistently elevated aminotransferase levels may persist despite HCV eradication. Aim: To investigate prevalence and risk factors for ongoing hepatic inflammation after SVR in two large patient cohorts. Methods This post-hoc analysis was based on prospectively collected demographic and clinical data from 834 patients with SVR after HCV treatment with either PegIFN- or DAA-based treatment regimens from the PRAMA trial (n = 341) or patients treated at our outpatient clinic (n = 493). Results We observed an unexpected high prevalence of post-SVR inflammation, including patients who received novel IFN-free DAA-based therapies. Up to 10% of patients had ongoing elevation of aminotransferase levels and another 25% showed aminotransferase activity above the so-called healthy range. Several baseline factors were independently associated with post-SVR aminotransferase elevation. Among those, particularly male gender, advanced liver disease and markers for liver steatosis were strongly predictive for persistent ALT elevation. The use of IFN-based antiviral treatment was independently correlated with post-SVR inflammation, further supporting the overall benefit of IFN-free combination regimens. Conclusion This is the first comprehensive study on a large patient cohort investigating the prevalence and risk factors for ongoing liver inflammation after eradication of HCV. Our data show a high proportion of patients with ongoing hepatic inflammation despite HCV eradication with potential implications for the management of approximately one third of all patients upon SVR. PMID:28196130

  13. Evaluation of hepatoprotective effect of methanolic extract of Clitoria ternatea (Linn.) flower against acetaminophen-induced liver damage

    PubMed Central

    Nithianantham, Kuppan; Ping, Kwan Yuet; Latha, Lachimanan Yoga; Jothy, Subramanion L; Darah, Ibrahim; Chen, Yeng; Chew, Ai-Lan; Sasidharan, Sreenivasan

    2013-01-01

    Objective To evaluate the hepatoprotective and antioxidant activity of Clitoria ternatea (C. ternatea) flower extract against acetaminophen-induced liver toxicity. Methods The antioxidant property of C. ternatea flower extract was investigated by employing established in vitro antioxidant assay. The C. ternatea flower extract was studied in this work for its hepatoprotective effect against acetaminophen-induced liver toxicity in mice. Activity was measured by monitoring the levels of aspartate aminotransferase, alanine aminotransferase, billirubin and glutathione with histopathological analysis. Results The amount of total phenolics and flavonoids were estimated to be 105.40±2.47 mg/g gallic acid equivalent and 72.21±0.05 mg/g catechin equivalent respectively. The antioxidant activity of C. ternatea flower extract was 68.9% at a concentration of 1 mg/mL and was also concentration dependant, with an IC50 value of 327.00 µg/mL. The results of acetaminophen-induced liver toxicity experiment showed that mice treated with the extract (200 mg/kg) showed a significant decrease in alanine aminotransferase, aspartate aminotransferase, and bilirubin levels, which were all elevated in the paracetamol group (P<0.05). Meanwhile, the level of glutathione was found to be restored in extract treated animals compared to the groups treated with acetaminophen alone (P<0.05). Therapy of extract also showed its protective effect on histopathological alterations and supported the biochemical finding. Conclusion The present work confirmed the hepatoprotective effect of C. ternatea flower against model hepatotoxicant acetaminophen.

  14. Weights, hematology and serum chemistry of free-ranging brown boobies (Sula leucogaster) in Johnston Atoll, Central Pacific

    USGS Publications Warehouse

    Work, T.M.

    1999-01-01

    Hematologic and serum chemistry values are reported for 105 brown boobies (Sula leucogaster) from Johnston Atoll, Central Pacific. Hematocrit, estimated total plasma solids, total and differential white cell counts, serum glucose, calcium, phosphorus, uric acid, total protein, albumin, globulin, aspartate aminotransferase, and creatinine phosphokinase were analyzed. Hematologic and serum chemistry values varied with age and sex. Values were compared with those of red-footed boobies and other tropical and temperate marine pelecaniforms.

  15. 17β-estradiol attenuates reduced-size hepatic ischemia/reperfusion injury by inhibition apoptosis via mitochondrial pathway in rats.

    PubMed

    Lin, Fu Sheng; Shen, Shi Qiang; Chen, Zu Bing; Yan, Rui Chen

    2012-02-01

    The aim of this study was to investigate the effect of 17β-estradiol (E2) on hepatocyte apoptosis after reduced-size hepatic ischemia/reperfusion (I/R) injury and its mechanism. A rat model of reduced-size hepatic I/R injury was established. Sprague-Dawley rats were randomly allocated into sham, I/R, and E2 + I/R group. 17β-Estradiol (4 mg/kg) or the vehicle was administered i.p. 1 h before ischemia and immediately after operation. For each group, 10 rats were used to investigate the survival during a week after reperfusion. Blood samples and liver tissues were obtained in the remaining animals after 3, 6, 12, and 24 h of reperfusion to assess serum aspartate aminotransferase and alanine aminotransferase levels, liver tissue malondialdehyde concentration, superoxide dismutase activity, and histopathologic changes. Apoptosis ratio; expression of cytochrome c, Bcl-2, and Bax proteins; and enzymatic activities of caspase 9 and caspase 3 were performed in the samples at 12 h after reperfusion. The serum aspartate aminotransferase and alanine aminotransferase levels and tissue malondialdehyde concentration were increased in the I/R group, whereas the increase was significantly reduced by E2. The superoxide dismutase activity, depressed by I/R injury, was elevated back to normal levels by treatment with E2. Severe hepatic damage was observed by light microscopy in the I/R group, whereas administration of E2 resulted in tissue and cellular preservation. Furthermore, E2 inhibited hepatocellular apoptosis by upregulating the ratio of Bcl-2 and Bax expression, reduced cytosolic cytochrome c level, and decreased caspase 9 and caspase 3 activities. The 7-day survival rate was significantly higher in the E2 + I/R group than in the I/R group. These results indicated that E2 protects liver tissues from reduced-size hepatic I/R injury by suppressing mitochondrial apoptotic pathways.

  16. Choledocholithiasis presenting with very high transaminase level

    PubMed Central

    Agahi, Amy; McNair, Alistair

    2012-01-01

    We present three cases of choledocholithiasis presenting with a rise in transaminase to levels normally associated with acute hepatitis (alanine aminotransferase in excess of 1000 IU/l). All three cases had repeated investigation for liver disease before identification of common bile duct stones with magnetic resonance cholangiopancreatogram, and removal at endoscopic retrograde cholangiopancreatogram. We discuss the existing literature and the potential mechanisms of hepatocyte injury in extrahepatic obstruction. PMID:23188856

  17. Preliminary Assessment of the Relative Toxicity of 1,5-Diazido-3- Nitrazapentane 90-Day Dermal Application Male and Female Rabbits Study Number 75-51-Y809-90, May 1990 - June 1992. Phase 2

    DTIC Science & Technology

    1992-06-01

    blood samples were collected by the cephalic ear artery from both male and female rabbits each dose group. Tables 4 and 5 list the clinical chemistry and...4. CLINICAL CHEMISTRY Serum Glutamic Oxaloacetic Transaminase (SGOT) (AST) (Aspartate Aminotransferase) Serum Glutamic Pyruvic Transaminase (SGPT...Appendices C-L). (3) The clinical chemistry values (Appendices I and J) showed a significant change in total protein in the 78 mg/kg group for male rabbits and

  18. Connection between hyperemesis gravidarum, jaundice or liver dysfunction, and biliary sludge.

    PubMed

    Matsubara, Shigeki; Kuwata, Tomoyuki; Kamozawa, Chihiro; Sakamoto, Yuki; Suzuki, Mitsuaki; Tamada, Kiichi

    2012-02-01

    Jaundice in hyperemesis gravidarum may cause physicians to suspect several underlying diseases. Jaundice appeared in a woman with hyperemesis gravidarum and an ultrasound revealed biliary sludge. Hydration concomitantly ameliorated the symptoms, jaundice and the biliary sludge. Another woman with hyperemesis gravidarum showed elevated aminotransferases, with biliary sludge also being present. Hydration ameliorated the symptoms and liver dysfunction, and reduced the total bilirubin level. Biliary sludge appeared, but was ameliorated according to the symptoms of hyperemesis gravidarum.

  19. A New Octadecenoic Acid Derivative from Caesalpinia gilliesii Flowers with Potent Hepatoprotective Activity

    PubMed Central

    Osman, Samir M.; El-Haddad, Alaadin E.; El-Raey, Mohamed A.; Abd El-Khalik, Soad M.; Koheil, Mahmoud A.; Wink, Michael

    2016-01-01

    Background: Caesalpinia gilliesii Hook is an ornamental shrub with showy yellow flowers. It was used in folk medicine due to its contents of different classes of secondary metabolites. In our previous study, dichloromethane extract of C. gilliesii flowers showed a good antioxidant activity. Aim of the Study: Isolation and identification of bioactive hepatoprotective compounds from C. gilliesii flowers dichloromethane fraction. Materials and Methods: The hepatoprotective activity of dichloromethane fraction and isolated compounds were studied in CCl4-intoxicated rat liver slices by measuring liver injury markers (alanine aminotransferase, aspartate aminotransferase and glutathione [GSH]). All compounds were structurally elucidated on the basis of electron ionization-mass spectrometry, one- and two-dimensional nuclear magnetic resonance. Results: A new 12,13,16-trihydroxy-14(Z)-octadecenoic acid was identified in addition to the known β-sitosterol-3-O-butyl, daucosterol, isorhamnetin, isorhamnetin-3-O-rhamnoside, luteolin-7,4’-dimethyl ether, genistein-5-methyl ether, luteolin-7-O-rhamnoside, isovanillic acid, and p-methoxybenzoic acid. Dichloromethane fraction and isorhamnetin were able to significantly protect the liver against intoxication. Moreover, the dichloromethane fraction and the isolated phytosterols induced GSH above the normal level. Conclusion: The hepatoprotective activity of C. gilliesii may be attributed to its high content of phytosterols and phenolic compounds. SUMMARY Bioactive Hepatoprotective phytosterols and phenolics from chloroform extract of Caesalpinia gilliesii Abbreviations used: ALT: Alanine Aminotransferase; AST: Aspartate aminotransferase; GSH: Glutathione; SC50: Scavenging Capacity 50 (SC 50); COSY: Correlation spectroscopy; NMR: Nuclear Magnetic Resonance; CC: Column chromatography; EI-MS: Electron-impact mass spectrometry; HSQC: Heteronuclear single-quantum correlation. PMID:27563221

  20. Pregravid Liver Enzyme Levels and Risk of Gestational Diabetes Mellitus During a Subsequent Pregnancy

    PubMed Central

    Xu, Fei; Darbinian, Jeanne; Quesenberry, Charles P.; Ferrara, Assiamira; Hedderson, Monique M.

    2014-01-01

    OBJECTIVE Liver enzymes are independent predictors of type 2 diabetes. Although liver fat content correlates with features of insulin resistance, a risk factor for developing gestational diabetes mellitus (GDM), the relationship between liver enzymes and GDM is unclear. The objective of this study was to assess whether pregravid liver enzyme levels are associated with subsequent risk of GDM. RESEARCH DESIGN AND METHODS A nested case-control study was conducted among women who participated in the Kaiser Permanente Northern California multiphasic health checkup (1984–1996) and had a subsequent pregnancy (1984–2009). Case patients were 256 women who developed GDM. Two control subjects were selected for each case patient and matched for year of blood draw, age at examination, age at pregnancy, and number of intervening pregnancies. RESULTS Being in the highest quartile versus the lowest quartile of γ-glutamyl transferase (GGT) levels was associated with a twofold increased risk of subsequent GDM (odds ratio 1.97 [95% CI 1.14–3.42]), after adjusting for race/ethnicity, prepregnancy BMI, family history of diabetes, and alcohol use. This result was attenuated after adjusting for homeostasis model assessment of insulin resistance (HOMA-IR), fasting status, and rate of gestational weight gain. There was significant interaction between GGT and HOMA-IR; the association with GGT was found among women in the highest tertile of HOMA-IR. Aspartate aminotransferase and alanine aminotransferase were not associated with increased GDM risk. CONCLUSIONS Pregravid GGT level, but not alanine aminotransferase or aspartate aminotransferase level, predicted the subsequent risk of GDM. Markers of liver fat accumulation, such as GGT level, are present years before pregnancy and may help to identify women at increased risk for subsequent GDM. PMID:24795397

  1. Physico-chemical and biochemical approaches to assessing the development of precancerous pathologies of the gastrointestinal tract during their modeling in mice with complex effects of stress factors of different nature

    NASA Astrophysics Data System (ADS)

    Alieva, Zamira O.; Kanevskiy, Matvej V.; Galitskaya, Anna A.; Mironova, Irina K.; Pleshakova, Ekaterina V.; Velikov, Vladimir A.; Chumakov, Daniil S.; Semyachkina-Glushkovskaya, Oxana V.; Konnova, Svetlana A.

    2016-04-01

    It was shown in the experience with 60 white mice that separate and combined effects of stress factors: noctidial lighting (800 lux), sodium nitrite (0.2% solution in water) and p-toluidine (with food) within 107 days causes a change in impedance value of erythrocytes at frequencies 1 Hz - 1MHz. Changes in the activity of intracellular aminotransferases, creatinine and urea were observed, indicating cardiotoxic and hepatotoxic effects.

  2. Hepatoprotective Effects of Corilagin Following Hemorrhagic Shock are Through Akt-Dependent Pathway.

    PubMed

    Liu, Fu-Chao; Chaudry, Irshad H; Yu, Huang-Ping

    2017-03-01

    Corilagin, a component of Phyllanthus urinaria extract, possesses antioxidant, thrombolytic, antiatherogenic, and hepatoprotective properties, but the mechanism underlying these effects remains unclear. Previous studies showed that the Akt (protein kinase B) signaling pathway exerts anti-inflammatory and organ protective effects. The aim of this study was to investigate the mechanism of action of corilagin and determine whether these effects are mediated through the Akt-dependent pathway in a trauma-hemorrhagic shock-induced liver injury rodent model. Hemorrhagic shock was induced in male Sprague-Dawley rats; mean blood pressure was maintained at 35 mm Hg to 40 mm Hg for 90 min, followed by fluid resuscitation. During resuscitation, three doses of corilagin alone (1 mg/kg, 5 mg/kg, or 10 mg/kg, intravenously) were administered. Furthermore, a single dose of corilagin (5 mg/kg) with and without Wortmannin (1 mg/kg, PI3K inhibitor), Wortmannin alone, or vehicle was administered. Twenty-four hours after resuscitation, plasma alanine aminotransferase and aspartate aminotransferase concentration and hepatic parameters were measured. One-way ANOVA was used for statistical analysis. Hepatic myeloperoxidase activity and the concentrations of plasma alanine aminotransferase and aspartate aminotransferase, interleukin-6, tumor necrosis factor-α, intercellular adhesion molecule-1, and cytokine-induced neutrophil chemoattractant-1 (CINC-1) and CINC-3 increased following hemorrhagic shock. These parameters were significantly attenuated in corilagin-treated rats following hemorrhagic shock. Hepatic phospho-Akt expression was also higher in corilagin-treated rats than in vehicle-treated rats. The elevation of phospho-Akt was abolished by combined treatment with Wortmannin and corilagin. Our results suggest that corilagin exerts its protective effects on hemorrhagic shock-induced liver injury, at least, via the Akt-dependent pathway.

  3. The study of some haematological and serum biochemical parameters of juvenile beluga (Huso huso) fed oligofructose.

    PubMed

    Hoseinifar, Seyed Hossein; Mirvaghefi, Alireza; Merrifield, Daniel L; Amiri, Bagher Mojazi; Yelghi, Saeed; Bastami, Kazem Darvish

    2011-03-01

    A study was conducted to investigate the effects of dietary oligofructose (1, 2 and 3%) on the blood profiles of beluga (Huso huso) juveniles (18.77 ± 0.76 g) compared to fish fed an un-supplemented diet. After 7 weeks of feeding on the experimental diets, haematological parameters, metabolic products (cholesterol, glucose and total protein) and serum enzymes (lactate dehydrogenase, alkaline phosphatase, alanine aminotransferase and aspartate aminotransferase) were measured. Compared to the control group (0% oligofructose), dietary oligofructose had no effect on red blood cell counts (RBC), mean corpuscular volume (MCV), mean cellular haemoglobin (MCH) or mean cell haemoglobin concentration (MCHC) (P > 0.05). However, haemoglobin (Hb) concentration, leucocyte (WBC) levels and the proportion of lymphocytes were significantly higher (P > 0.05) in the 2% oligofructose fed fish than in the 3% oligofructose fed fish. Additionally, haematocrit (Hct) values (P = 0.049) and the proportion of lymphocytes (P ≤ 0.01) were significantly higher in the 2% oligofructose group than in the control group. Although serum glucose and total protein remained unaffected, serum cholesterol was significantly lower in the 2% oligofructose group than in the control and 3% oligofructose group (P < 0.05). The results of the present study showed that oligofructose had no significant effects on serum lactate dehydrogenase, alkaline phosphatase, alanine aminotransferase and aspartate aminotransferase. These results indicate that fish blood profiles could be affected by prebiotics, which should be taken into account in future studies.

  4. Rhabdomyolysis and acute kidney injury after acupuncture sessions.

    PubMed

    Papasotiriou, Marios; Betsi, Grigoria; Tsironi, Maria; Assimakopoulos, Georgios

    2014-05-01

    Rhabdomyolysis is usually caused by muscle injury, drugs or alcohol and presents with muscle weakness and pain. It is characterized by rise in serum creatine kinase, aminotransferases and electrolytes as well as myoglobinuria. Myoglobinuria may cause acute kidney injury by direct proximal tubule cytotoxicity, renal vasoconstriction, intraluminal cast formation and distal tubule obstruction. Muscle pain and weakness as well as vascular injury have been reported after acupuncture. We report a case of severe rhabdomyolysis and acute kidney injury after acupuncture sessions.

  5. Development and testing of a contamination potential mapping system for a portion of the General Separations Area, Savannah River Site, South Carolina

    USGS Publications Warehouse

    Rine, J.M.; Berg, R.C.; Shafer, J.M.; Covington, E.R.; Reed, J.K.; Bennett, C.B.; Trudnak, J.E.

    1998-01-01

    A methodology was developed to evaluate and map the contamination potential or aquifer sensitivity of the upper groundwater flow system of a portion of the General Separations Area (GSA) at the Department of Energy's Savannah River Site (SRS) in South Carolina. A Geographic Information System (GIS) was used to integrate diverse subsurface geologic data, soils data, and hydrology utilizing a stack-unit mapping approach to construct mapping layers. This is the first time that such an approach has been used to delineate the hydrogeology of a coastal plain environment. Unit surface elevation maps were constructed for the tops of six Tertiary units derived from over 200 boring logs. Thickness or isopach maps were created for five hydrogeologic units by differencing top and basal surface elevations. The geologic stack-unit map was created by stacking the five isopach maps and adding codes for each stack-unit polygon. Stacked-units were rated according to their hydrogeologic properties and ranked using a logarithmic approach (utility theory) to establish a contamination potential index. Colors were assigned to help display relative importance of stacked-units in preventing or promoting transport of contaminants. The sensitivity assessment included the effects of surface soils on contaminants which are particularly important for evaluating potential effects from surface spills. Hydrogeologic/hydrologic factors did not exhibit sufficient spatial variation to warrant incorporation into contamination potential assessment. Development of this contamination potential mapping system provides a useful tool for site planners, environmental scientists, and regulatory agencies.A methodology was developed to evaluate and map the contamination potential or aquifer sensitivity of the upper groundwater flow system of a portion of the General Separations Area (GSA) at the Department of Energy's Savannah River Site (SRS) in South Carolina. A Geographic Information System (GIS) was used to

  6. Electrochemical biosensor array for liver diagnosis using silanization technique on nanoporous silicon electrode.

    PubMed

    Song, Min-Jung; Yun, Dong-Hwa; Min, Nam-Ki; Hong, Suk-In

    2007-01-01

    An electrochemical biosensor array system was fabricated for the diagnosis and monitoring of liver diseases. Analysis on this array system with multiple samples was performed for point-of-care testing or home-use applications. Cholesterol, bilirubin and aminotransferases present in the serum are well-known biomarkers for liver diseases. For this study, we describe our biosensor array system consisting of cholesterol, bilirubin and glutamate sensors. To immobilize sensing enzymes on the array system, we employed a silanization technique. We observed that porous silicon layers formed on each working electrode notably increase the effective surface area. Sensing electrodes were placed in sampling wells to minimize the cross-interference effect so that multiple sampling would be possible with a low noise current. Compared with traditional analyte measurement procedures, our novel analytical device demonstrated acceptable sensitivities for the analyses of multiple samples and analytes without a marked cross-interference effect. The device sensitivities observed were 0.2656 microA/mM for cholesterol, 0.15354 mA/mM for bilirubin, 0.13698 microA/(U/l) for alanine aminotransferase (ALT) and 0.45439 microA/(U/l) for aspartate aminotransferase (AST).

  7. Hematological and plasma biochemical reference ranges of Alaskan seabirds: Their ecological significance and clinical importance

    USGS Publications Warehouse

    Newman, S.H.; Piatt, J.F.; White, J.

    1997-01-01

    Blood was analyzed from 151 pelagic marine birds to establish reference ranges for hematological and plasma biochemical parameters from healthy, wild populations of Pacific seabirds. Of the 13 species examined, 9 were from the Family Alcidae (N = 122 individuals) and the remainder (N = 29) from the Families Phalacrocoracidae, Laridae, and Procellariidae. Three of 8 hematological parameters (total white blood cell count, lymphocyte count and eosinophil count) differed significantly among species, as did 9 of 13 plasma biochemical parameters (alkaline phosphatase, aspartate aminotransferase, creatine kinase, cholesterol, glucose, lactate dehydrogenase, total bilirubin, total protein and field total protein). There were no differences among species for packed cell volume, buffy coat, cell counts of heterophils, monoqtes and basophils, or for concentrations of alanine aminotransferase, triglycerides, uric acid and calcium. Plasma calcium concentration, triglyceride levels and field total protein varied significantly between sexes, with females having higher mean concentrations of all 3 parameters. However, no significant relationships between measures of breeding condition (brood patch size, subcutaneous and mesenteric fat deposits, or ovarian follicle size and ovary weight) and calcium or alkaline phosphatase concentrations in female birds could be identified. Alanine aminotransferase and uric acid were the only analytes which did not differ significantly between species or sexes.

  8. Artemether Exhibits Amoebicidal Activity against Acanthamoeba castellanii through Inhibition of the Serine Biosynthesis Pathway

    PubMed Central

    Deng, Yihong; Ran, Wei; Man, Suqin; Li, Xueping; Gao, Hongjian; Tang, Wei

    2015-01-01

    Acanthamoeba sp. parasites are the causative agents of Acanthamoeba keratitis, fatal granulomatous amoebic encephalitis, and cutaneous infections. However, there are currently no effective drugs for these organisms. Here, we evaluated the activity of the antimalarial agent artemether against Acanthamoeba castellanii trophozoites and identified potential targets of this agent through a proteomic approach. Artemether exhibited in vitro amoebicidal activity in a time- and dose-dependent manner and induced ultrastructural modification and cell apoptosis. The iTRAQ quantitative proteomic analysis identified 707 proteins that were differentially expressed after artemether treatment. We focused on phosphoglycerate dehydrogenase and phosphoserine aminotransferase in the serine biosynthesis pathway because of their importance to the growth and proliferation of protozoan and cancer cells. The expression of these proteins in Acanthamoeba was validated using quantitative real-time PCR and Western blotting after artemether treatment. The changes in the expression levels of phosphoserine aminotransferase were consistent with those of phosphoglycerate dehydrogenase. Therefore, the downregulation of phosphoserine aminotransferase may be due to the downregulation of phosphoglycerate dehydrogenase. Furthermore, exogenous serine might antagonize the activity of artemether against Acanthamoeba trophozoites. These results indicate that the serine biosynthesis pathway is important to amoeba survival and that targeting these enzymes would improve the treatment of Acanthamoeba infections. Artemether may be used as a phosphoglycerate dehydrogenase inhibitor to control or block Acanthamoeba infections. PMID:26014935

  9. Hepatoprotective effects of Spirulina maxima in patients with non-alcoholic fatty liver disease: a case series

    PubMed Central

    2010-01-01

    Introduction Non-alcoholic fatty liver diseases range from simple steatosis to non-alcoholic steatohepatitis. The "two hits" hypothesis is widely accepted for its pathogenesis: the first hit is an increased fat flux to the liver, which predisposes our patient to a second hit where increasing free fatty acid oxidation into the mitochondria leads to oxidative stress, lipoperoxidation and a chain reaction with increased ROS. Clinical indications include abdominal cramps, meteorism and fatigue. Most patients, however, are asymptomatic, and diagnosis is based on aminotransferase elevation and ultrasonography (or "brilliant liver"). Spirulina maxima has been experimentally proven to possess in vivo and in vitro hepatoprotective properties by maintaining the liver lipid profile. This case report evaluates the hepatoprotective effects of orally supplied Spirulina maxima. Case presentation Three Hispanic Mexican patients (a 43-year-old man, a 77-year-old man and a 44-year-old woman) underwent ultrasonography and were treated with 4.5 g/day of Spirulina maxima for three months. Their blood samples before and after the treatment determined triacylglycerols, total cholesterol, high-density lipoprotein cholesterol, alanine aminotransferase and low-density lipoprotein cholesterol levels. The results were assessed using ultrasound. Conclusion Treatment had therapeutic effects as evidenced by ultrasonography and the aminotransferase data. Hypolipidemic effects were also shown. We conclude that Spirulina maxima may be considered an alternative treatment for patients with non-alcoholic fatty liver diseases and dyslipidemic disorder. PMID:20370930

  10. [Dopamine content in blood and activity of alcohol-transforming enzymes in alcoholism].

    PubMed

    Kharchenko, N K

    1997-01-01

    An increase of alcohol dehydrogenase activity is observed in patients with chronic alcoholism at the first stage of the disease under normal indices of activity of aldehyde dehydrogenase, aspartate- and alanine aminotransferase and thymol sample that evidences for the induction of alcohol dehydrogenase synthesis in the liver. At the second stage of alcoholism the activity of alcohol dehydrogenase, aspartate- and alanine aminotransferase, the index of thymol sample increase while activity of aldehyde dehydrogenase decreases that indicates to organic destructive changes in the liver. At the third stage of alcoholism one can observe the decrease in activity of alcohol dehydrogenase, aldehyde dehydrogenase and alanine aminotransferase relative to activity of these enzymes at the second stage, that can evidence for the increase of the possibility of the processes of synthesis of the liver. The correlation of alcohol dehydrogenase activity to that of aldehyde dehydrogenase in the process of formation and development of alcoholism is shifted towards the progressive accumulation of acetaldehyde. Parallel increase of dopamine concentration in blood creates conditions for formation of morphine-like alcaloides--products of condensation of acetaldehide with dopamine.

  11. Cytomegalovirus hepatitis and myopericarditis.

    PubMed

    Zubiaurre, Leire; Zapata, Eva; Bujanda, Luis; Castillo, María; Oyarzabal, Igor; Gutiérrez-Stampa, Maria A; Cosme, Angel

    2007-01-28

    Cytomegalovirus (CMV) infection in inmunocompetent hosts generally is asymptomatic or may present as a mononucleosis syndrome but rarely can lead to severe organ complications. We report a case of simultaneous hepatic and pericardic CMV infection in a 36-year old immunocompetent man. He was admitted to coronary unit with fever, chest pain radiated to shoulders, changes on electrocardiogram with diffuse ST elevation and modest laboratory elevations in the MB fraction of creatine kinase (CK-MB) of 33.77 microg/L (0.1-6.73), serum cardiac troponin T of 0.904 ng/mL (0-0.4), creatine kinase of 454 U/L (20-195) and myoglobin of 480.4 microg/L (28-72). Routine laboratory test detected an elevation of aminotransferase level: alanine aminotransferase 1445 U/L, aspartate aminotransferase 601 U/L. We ruled out other causes of hepatitis with normal results except IgM CMV. The patient was diagnosed with myopericarditis and hepatitis caused by cytomegalovirus and started symptomatic treatment with salicylic acid. In few days the laboratory findings became normal and the patient was discharged.

  12. Isolation of Intact Chloroplasts and Other Cell Organelles from Spinach Leaf Protoplasts 1

    PubMed Central

    Nishimura, Mikio; Graham, Douglas; Akazawa, Takashi

    1976-01-01

    Freshly prepared spinach leaf protoplasts were gently ruptured by mechanical shearing followed by sucrose density gradient centrifugation to separate constituent cell organelles. The isolation of intact Class I chloroplasts (d = 1.21) in high yield, well separated from peroxisomes and mitochondria, was evidenced by the specific localization of ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39), NADP triose-P dehydrogenase (EC 1.2.1.9), and carbonic anhydrase (EC 4.2.1.1) in the fractions. A clear separation of chloroplastic ribosomes from the soluble cytoplasmic ribosomes was also demonstrated by the band patterns of constituent RNA species in the polyacrylamide gel electrophoresis. Localization of several enzyme activities specific to leaf peroxisomes, e.g. catalase (EC 1.11.1.6), glycolate oxidase (EC 1.1.3.1), glyoxylate reductase (EC 1.1.1.26), glutamate glyoxylate aminotransferase (EC 2.6.1.4), serine glyoxylate aminotransferase, and alanine glyoxylate aminotransferase (EC 2.6.1.12) in the peroxisomal fractions (d = 1.25), was demonstrated. Overall results show the feasibility of the method for the isolation of pure organelle components in leaf tissues. Images PMID:16659669

  13. Physiological and biochemical parameters in response to electroejaculation in adult and yearling anesthetized pampas deer (Ozotoceros bezoarticus) males.

    PubMed

    Fumagalli, F; Villagrán, M; Damián, J P; Ungerfeld, R

    2012-04-01

    The aim of this study was to characterize changes in physiological and biochemical parameters during electroejaculation (EE) under general anaesthesia in adult and yearling pampas deer males (Ozotoceros bezoarticus). The relation between heart rate, pulse rate, respiratory rate and oximetry with EE voltages was studied. The changes in cortisol, alkaline phosphatase, creatine kinase, aspartate aminotransferase and rectal temperature were determined before and after electroejaculation (BEE and AEE). Heart rate and pulse rate values increased during EE, with a greater increase in heart rate in adults (p < 0.01). Respiratory rate and SpO(2) were not affected by EE or category. The rectal temperature decreased in adults and yearlings during EE (p = 0.0001). Alkaline phosphatase, creatine kinase and aspartate aminotransferase concentrations increased AEE (p < 0.001, p < 0.01 and p < 0.05, respectively). Alkaline phosphatase and creatine kinase concentrations were greater in young than in adult males (p < 0.05). In conclusion, the EE under general anaesthesia in the pampas deer provoked increases in heart and pulse rates, as well as in creatine kinase, aspartate aminotransferase and alkaline phosphatase concentrations, and a decrease in rectal temperature in both categories. Oximetry values were especially low during the procedure. Heart rate increase was greater in adult males, and alkaline phosphatase and creatine kinase concentrations were greater in yearlings.

  14. Hypolipidemic effect of the edible mushroom Agaricus blazei in rats subjected to a hypercholesterolemic diet.

    PubMed

    de Miranda, Aline M; Ribeiro, Gustavo M; Cunha, Aureliano C; Silva, Lorena S; dos Santos, Rinaldo C; Pedrosa, Maria Lúcia; Silva, Marcelo E

    2014-03-01

    The effects of Agaricus blazei intake on the lipid profile of animals fed a hypercholesterolemic diet were evaluated. Thirty-two female Fisher rats were divided into four groups and given the standard AIN-93 M diet (C), this diet + 1 % A. blazei (CAb), a hypercholesterolemic diet with 25 % soybean oil and 1 % cholesterol (H) or this diet + 1 % A. blazei (HAb) for 6 weeks. Food intake, weight gain, liver and serum lipid profiles, activity of aminotransferases [alanine aminotransferase (ALT) and aspartate aminotransferase (AST)], and creatinine and urea levels as well as abdominal fat weight were measured. Histological analysis of kidney and liver tissue was also performed. The HAb group had a higher food intake, but a lower weight gain as compared to group H. This resulted in a significant decrease in abdominal fat weight, to values close to those of groups C and CAb. Supplementing the hypercholesterolemic diet with A. blazei promoted a significant reduction in total and non-HDL cholesterol, as well as in the atherogenic index, as compared to group H, and this effect was more pronounced in the serum. There was no hepatotoxic effect caused by the supplementation of the diets with the mushroom. We conclude that in our experimental model and in the concentration used, A. blazei was effective in improving the lipid profile of the animals.

  15. Artemether Exhibits Amoebicidal Activity against Acanthamoeba castellanii through Inhibition of the Serine Biosynthesis Pathway.

    PubMed

    Deng, Yihong; Ran, Wei; Man, Suqin; Li, Xueping; Gao, Hongjian; Tang, Wei; Tachibana, Hiroshi; Cheng, Xunjia

    2015-08-01

    Acanthamoeba sp. parasites are the causative agents of Acanthamoeba keratitis, fatal granulomatous amoebic encephalitis, and cutaneous infections. However, there are currently no effective drugs for these organisms. Here, we evaluated the activity of the antimalarial agent artemether against Acanthamoeba castellanii trophozoites and identified potential targets of this agent through a proteomic approach. Artemether exhibited in vitro amoebicidal activity in a time- and dose-dependent manner and induced ultrastructural modification and cell apoptosis. The iTRAQ quantitative proteomic analysis identified 707 proteins that were differentially expressed after artemether treatment. We focused on phosphoglycerate dehydrogenase and phosphoserine aminotransferase in the serine biosynthesis pathway because of their importance to the growth and proliferation of protozoan and cancer cells. The expression of these proteins in Acanthamoeba was validated using quantitative real-time PCR and Western blotting after artemether treatment. The changes in the expression levels of phosphoserine aminotransferase were consistent with those of phosphoglycerate dehydrogenase. Therefore, the downregulation of phosphoserine aminotransferase may be due to the downregulation of phosphoglycerate dehydrogenase. Furthermore, exogenous serine might antagonize the activity of artemether against Acanthamoeba trophozoites. These results indicate that the serine biosynthesis pathway is important to amoeba survival and that targeting these enzymes would improve the treatment of Acanthamoeba infections. Artemether may be used as a phosphoglycerate dehydrogenase inhibitor to control or block Acanthamoeba infections.

  16. Hepatoprotective activity of quercetin against acrylonitrile-induced hepatotoxicity in rats.

    PubMed

    Abo-Salem, Osama M; Abd-Ellah, Mohamed F; Ghonaim, Mabrouk M

    2011-01-01

    Acrylonitrile is a potent hepatotoxic, mutagen, and carcinogen. A role for free radical-mediated lipid peroxidation in the toxicity of acrylonitrile has been suggested. The present study was designed to assess the hepatoprotective effect of quercetin against acrylonitrile-induced hepatotoxicity in rats. Liver damage was induced by oral administration of acrylonitrile (50 mg/kg/day/5 weeks). Acrylonitrile produced a significant elevation of malondialdehyde (138.9%) with a marked decrease in reduced glutathione (72.4%), and enzymatic antioxidants; superoxide dismutase (81%), and glutathione peroxidase (53.2%) in the liver. Serum aspartate aminotransferase, alanine aminotransferases, direct bilirubin, and total bilirubin showed a significant increase in acrylonitrile alone treated rats (115.5%, 110.8%, 1006.8%, and 1000.8%, respectively). Pretreatment with quercetin (70 mg/kg/day/6 weeks) and its coadministration with acrylonitrile prevented acrylonitrile-induced alterations in hepatic lipid peroxides and enzymatic antioxidants as well as serum aminotransferases and bilirubin. Histopathological findings supported the biochemical results. We suggest that querectin possess hepatoprotective effect against acrylonitrile-induced hepatotoxicity through its antioxidant activity.

  17. IFCC primary reference procedures for the measurement of catalytic activity concentrations of enzymes at 37 °C. Part 9: reference procedure for the measurement of catalytic concentration of alkaline phosphatase International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) Scientific Division, Committee on Reference Systems of Enzymes (C-RSE) (1)).

    PubMed

    Schumann, Gerhard; Klauke, Rainer; Canalias, Francesca; Bossert-Reuther, Steffen; Franck, Paul F H; Gella, F-Javier; Jørgensen, Poul J; Kang, Dongchon; Lessinger, Jean-Marc; Panteghini, Mauro; Ceriotti, Ferruccio

    2011-09-01

    Abstract This paper is the ninth in a series dealing with reference procedures for the measurement of catalytic activity concentrations of enzymes at 37 °C and the certification of reference preparations. Other parts deal with: Part 1. The concept of reference procedures for the measurement of catalytic activity concentrations of enzymes; Part 2. Reference procedure for the measurement of catalytic concentration of creatine kinase; Part 3. Reference procedure for the measurement of catalytic concentration of lactate dehydrogenase; Part 4. Reference procedure for the measurement of catalytic concentration of alanine aminotransferase; Part 5. Reference procedure for the measurement of catalytic concentration of aspartate aminotransferase; Part 6. Reference procedure for the measurement of catalytic concentration of γ-glutamyltransferase; Part 7. Certification of four reference materials for the determination of enzymatic activity of γ-glutamyltransferase, lactate dehydrogenase, alanine aminotransferase and creatine kinase at 37 °C; Part 8. Reference procedure for the measurement of catalytic concentration of α-amylase. The procedure described here is derived from the previously described 30 °C IFCC reference method. Differences are tabulated and commented on in Appendix 1.

  18. Characterization of the enzymic capacity for cysteine desulphhydration in liver and kidney of the rat.

    PubMed Central

    Stipanuk, M H; Beck, P W

    1982-01-01

    The contribution of cystathionine gamma-lyase, cystathionine beta-synthase and cysteine aminotransferase coupled to 3-mercaptopyruvate sulphurtransferase to cysteine desulphhydration in rat liver and kidney was assessed with four different assay systems. Cystathionine gamma-lyase and cystathionine beta-synthase were active when homogenates were incubated with 280 mM-L-cysteine and 3 mM-pyridoxal 5'-phosphate at pH 7.8. Cysteine aminotransferase in combination with 3-mercaptopyruvate sulphurtransferase catalysed essentially all of the H2S production from cysteine at pH 9.7 with 160 mM-L-cysteine, 2 mM-pyridoxal 5'-phosphate, 3 mM-2-oxoglutarate and 3 mM-dithiothreitol. At more-physiological concentrations of cysteine (2 mM) cystathionine gamma-lyase and cystathionine beta-synthase both appeared to be active in cysteine desulphhydration, whereas the aminotransferase pathway did not. The effect of inhibition of cystathionine gamma-lyase by a suicide inactivator, propargylglycine, in the intact rat was also investigated; there was no significant effect of propargylglycine administration on the urinary excretion of total 35S, 35SO4(2-) or [35S]taurine formed from labelled dietary cysteine. PMID:7150244

  19. Application of clinical laboratory measurements to issues of environmental health.

    PubMed

    Rej, R; Silkworth, J B; DeCaprio, A P

    1992-03-13

    Monitoring of biochemical constituents in serum is an important component in revealing potential toxicity in humans and experimental animals due to exposure to a variety of xenobiotic agents. The relative toxicity of pure compounds, usually at large doses, has helped elucidate the mode of action of these compounds and their relative risk. However, most actual cases of environmental exposure present an extensive range of components and the potential for synergistic or inhibitory interactions. In this paper we review two such environmental cases: The Love Canal chemical dump site in Niagara Falls, NY, and the transformer fire at the State Office Building in Binghamton, NY. We focus on the clinical laboratory measurements obtained in these studies (including serum glucose, triglycerides, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, gamma-glutamyltransferase, lactate dehydrogenase, sodium and potassium), their usefulness, limitations, and application to such cases. Significant alterations in serum triglyceride and alanine aminotransferase levels were found in guinea pigs due to exposure to dioxins. These two tests were useful in estimating the 'equivalent' concentration of 2,3,7,8-tetrachlorodibenzo-p-dioxin in complex chemical mixtures.

  20. Effect of Inhaling Cymbopogon martinii Essential Oil and Geraniol on Serum Biochemistry Parameters and Oxidative Stress in Rats

    PubMed Central

    Andrade, Bruna Fernanda Murbach Teles; Braga, Camila Pereira; dos Santos, Klinsmann Carolo; Barbosa, Lidiane Nunes; Rall, Vera Lúcia Mores; Sforcin, José Maurício; Fernandes, Ana Angélica Henrique; Fernandes Júnior, Ary

    2014-01-01

    The effects of the inhalation of Cymbopogon martinii essential oil (EO) and geraniol on Wistar rats were evaluated for biochemical parameters and hepatic oxidative stress. Wistar rats were divided into three groups (n = 8): G1 was control group, treated with saline solution; G2 received geraniol; and G3 received C. martinii EO by inhalation during 30 days. No significant differences were observed in glycemia and triacylglycerol levels; G2 and G3 decreased (P < 0.05) total cholesterol level. There were no differences in serum protein, urea, aspartate aminotransferase activity, and total hepatic protein. Creatinine levels increased in G2 but decreased in G3. Alanine aminotransferase activity and lipid hydroperoxide were higher in G2 than in G3. Catalase and superoxide dismutase activities were higher in G3. C. martinii EO and geraniol increased glutathione peroxidase. Oxidative stress caused by geraniol may have triggered some degree of hepatic toxicity, as verified by the increase in serum creatinine and alanine aminotransferase. Therefore, the beneficial effects of EO on oxidative stress can prevent the toxicity in the liver. This proves possible interactions between geraniol and numerous chemical compounds present in C. martinii EO. PMID:25574396

  1. Indole-3-acetic Acid Synthesis in Tumorous and Nontumorous Species of Nicotiana 1

    PubMed Central

    Liu, Shih-Tung; Katz, Charles D.; Knight, C. Arthur

    1978-01-01

    The synthesis of indole-3-acetic acid (IAA) in the enzyme extracts of Nicotiana glauca, Nicotiana langsdorffii, their F1 hybrid, their amphidiploid hybrid, and the nontumorous mutant of the hybrid was investigated. Tryptamine, a possible precursor of IAA biosynthesis in Nicotiana tabacum, was not found in the callus tissue of N. glauca, N. langsdorffii, and their F1 hybrid. In petiole slices, the synthesis of IAA progressively increased during 5 hours of incubation in [14C]tryptophan. The rate of synthesis was about equal in the hybrid and N. langsdorffii but lower in N. glauca on either a cell or fresh weight basis. It was also found that tryptophan was about 25 times more efficient than tryptamine in promoting synthesis of IAA in petiole slices. It was found that indoleacetaldehyde oxidase, indoleacetaldehyde reductase, and tryptophan aminotransferase activities were present in all of the species examined; however, tryptophan decarboxylase activity was not found. The tryptophan aminotransferase activity in N. glauca, N. langsdorffii, and the nontumorous mutant required α-ketoglutaric acid and pyridoxal 5-phosphate whereas the addition of pyridoxal 5-phosphate seemed not to increase the enzyme activity in tumor plants. The tryptophan aminotransferase in the amphidiploid hybrid was partially purified by acetone precipitation. The enzyme activity had a temperature optimum at 49 C and a pH optimum at 8.9. It is suggested that there is an indolepyruvic acid pathway in the synthesis of IAA in the Nicotiana species examined. PMID:16660376

  2. Biochemical profile of Achatina fulica (Mollusca: Gastropoda) after infection by different parasitic loads of Angiostrongylus cantonensis (Nematoda, Metastrongylidae).

    PubMed

    Tunholi-Alves, Vinícius Menezes; Tunholi, Victor Menezes; Amaral, Ludimila Santos; Mota, Esther Maria; Maldonado Júnior, Arnaldo; Pinheiro, Jairo; Garcia, Juberlan

    2015-01-01

    The effect of experimental infection by different parasitic loads of Angiostrongylus cantonensis (Nematode, Metastrongylidae) on the activities of the aminotransferases and concentration of total proteins, uric acid and urea in the hemolymph of Achatina fulica (Mollusca, Gastropoda) were investigated. There was a significant decrease in the concentration of total proteins in the exposed snails to 5000 or more larvae. This change was accompanied by an increase in the concentrations of urea and uric acid in the hemolymph, suggesting a higher rate of deamination of the amino acids. Besides this, variations in the activities of the aminotransferases were also observed, with the highest values recorded in the groups exposed to greater parasite load. These results suggest an increase in the use of total proteins, since there was increased formation of nitrogenous catabolites, in conformity with an increase in the aminotransferase activities. Infection was verified by the fact that L3 larvae recovered from the snails was proportion to the exposure dose of L1 larvae. Histopathological results also indicated presence of an inflammatory cell infiltrate, favoring an increase of both transaminases.

  3. Effect of ketogenic diet on nucleotide hydrolysis and hepatic enzymes in blood serum of rats in a lithium-pilocarpine-induced status epilepticus.

    PubMed

    da Silveira, Vanessa Gass; de Paula Cognato, Giana; Müller, Alexandre Pastoris; Figueiró, Fabrício; Bonan, Carla Denise; Perry, Marcos L Santos; Battastini, Ana Maria Oliveira

    2010-06-01

    The ketogenic diet (KD) is a high-fat and low-carbohydrate diet, used for treating refractory epilepsy in children. We have previously shown alterations in nucleotidase activities from the central nervous system and blood serum of rats submitted to different models of epilepsy. In this study we investigated the effect of KD on nucleotidase activities in the blood serum, as well if KD has any influence in the activity of liver enzymes such as alkaline phosphatase, aspartate aminotransferase, and alanine aminotransferase activities in Wistar rats submitted to the lithium-pilocarpine model of epilepsy. At 21 days of age, rats received an injection of lithium chloride and, 18-19 h later, they received an injection of pilocarpine hydrochloride for status epilepticus induction. The results reported herein show that seizures induced by lithium-pilocarpine elicit a significant increase in ATP hydrolysis and alkaline phosphatase activity, as well as a decrease in ADP hydrolysis and aspartate aminotransferase activity. The KD is a rigorous regimen that can be associated with hepatic damage, as shown herein by the elevated activities of liver enzymes and 5'-nucleotidase in blood serum. Further studies are necessary to investigate the mechanism of inhibition of lithium on nucleotidases in blood serum.

  4. Polymorphisms and features of cytomegalovirus UL144 and UL146 in congenitally infected neonates with hepatic involvement.

    PubMed

    Guo, Gangqiang; Zhang, Liang; Ye, Sisi; Hu, Yingying; Li, Baoqing; Sun, Xiangwei; Mao, Chenchen; Xu, Jianfeng; Chen, Yiping; Zhang, Lifang; Xue, Xiangyang

    2017-01-01

    Human cytomegalovirus is a significant agent of hepatic involvement in neonates. In this study, we investigated the polymorphisms and features of the viral genes UL144 and UL146 as well as their significance to congenital hepatic involvement. In 79 neonates with congenital cytomegalovirus infection and hepatic involvement, full length UL144 and UL146 were successfully amplified in 73.42% and 60.76% of cases, respectively. Sequencing indicated that both genes were hypervariable. Notably, UL144 genotype B was highly associated with aspartate aminotransferase (P = 0.028) and lactate dehydrogenase (P = 0.046). Similarly, UL146 genotype G1 and G13 were significantly associated with CMV IgM (P = 0.026), CMV IgG (P = 0.034), alanine aminotransferase (P = 0.019), and aspartate aminotransferase (P = 0.032). In conclusion, dominant UL144 (genotype B) and UL146 (genotype G1 and G13) genotypes are associated with elevated levels of enzymes and CMV IgM and IgG of cytomegalovirus infection.

  5. Effects of lead shot ingestion on delta-aminolevulinic acid dehydratase activity, hemoglobin concentration, and serum chemistry in bald eagles

    USGS Publications Warehouse

    Hoffman, D.J.; Pattee, O.H.; Wiemeyer, Stanley N.; Mulhern, B.

    1981-01-01

    Lead shot ingestion by bald eagles (Haliaeetus leucocephalus) is considered to be widespread and has been implicated in the death of eagles in nature. It was recently demonstrated under experimental conditions that ingestion of as few as 10 lead shot resulted in death within 12 to 20 days. In the present study hematological responses to lead toxicity including red blood cell ALAD activity, hemoglobin concentration and 23 different blood serum chemistries were examined in five captive bald eagles that were unsuitable for rehabilitation and release. Eagles were dosed by force-feeding with 10 lead shot; they were redosed if regurgitation occurred. Red blood cell ALAD activity was inhibited by nearly 80% within 24 hours when mean blood lead concentration had increased to 0.8 parts per million (ppm). By the end of 1 week there was a significant decrease (20-25%) in hematocrit and hemoglobin, and the mean blood lead concentration was over 3 ppm. Within as little as 1-2 weeks after dosing, significant elevations in serum creatinine and serum alanine aminotransferase occurred, as well as a significant decrease in the ratio of serum aspartic aminotransferase to serum alanine aminotransferase. The mean blood lead concentration was over 5 ppm by the end of 2 weeks. These changes in serum chemistry may be indicative of kidney and liver alterations.

  6. New potential nonsteroidal anti-inflammatory drugs with antileukotrienic effects: influence on model proteins with catalytic activity.

    PubMed

    Netopilová, Miloslava; Drsata, Jaroslav; Beránek, Martin; Palicka, Vladimír

    2002-01-01

    Unspecific and side effects caused by interaction with proteins belong to common problems of many structures synthesized as potential medicaments. Possible in vitro interactions with proteins of a group of phenylsulfonyl benzoic acid derivatives (VUFB 19363, 19369, 19370, 19371, and 19760) as new potential anti-inflammatory compounds with anti-leukotrienic activities were studied in the present work. Three purified enzymes were used as model proteins with catalytic activities: Pig heart aspartate aminotransferase (AST, EC 2.6.1.1), alanine aminotransferase (ALT, EC 2.6.1.2), and glutamate decarboxylase (GAD, EC 4.1.1.15) from E. coli. Catalytic activities during incubation of individual compounds (6 x 10(-5) M solution to 5 x 10(-2) M suspension) at 37 degrees C with enzymes served as criteria of stability and function of the proteins. No immediate influence of any compound studied on enzyme activities was found. Aminotransferase activities were not affected even during incubation up to 20 d. In the case of GAD, the compounds VUFB 19369, 19370, 19371, and 19760 had stabilizing influence on GAD activity during incubation at enzyme concentrations of 11.25 and 5.62 mg prot/l. The lack of an immediate effect of compounds and the stability of enzymes during incubation them are favorable and support the prospective of the compounds as potential drugs.

  7. The effects of Artemisia aucheri extract on hepatotoxicity induced by thioacetamide in male rats

    PubMed Central

    Rezaei, Azam; ShekarForoush, Shahnaz; Changizi Ashtiyani, Saeed; Aqababa, Hydar; Zarei, Ali; Azizi, Maryam; Yarmahmodi, Hasan

    2013-01-01

    Objective: Liver is an important organ that is exposed to many oxidant and carcinogenic agents, thus antioxidant compounds are beneficial for liver health. Artemisia contains flavonoid compounds and anti-diabetic, antioxidant, and anti-inflammatory properties. Due to possessing terpene and sesquiterpene compounds, this plant has antioxidant properties. This study was done to investigate the effects of Artemisia plant extract on thioacetamide-induced hepatotoxicity in Wistar rats. Materials and Methods: For induction of hepatotoxicity, 50 mg/kg thioacetamide was injected intraperitoneally (i.p). After extraction and purification, the hydroalcoholic extract was injected i.p. at 100, 200, and 300 mg/kg doses for 21 days together with thioacetamide at 50 mg/kg dose in the last 3 days. After blood sampling and separation of serum, alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), albumin, and total protein concentrations were measured. Results: Significant decreases in aminotransferase and alkaline phosphatase activities and significant increases in the concentration of albumin and total protein in groups treated with the extract compared with thioacetamide-treated group were observed (p<0.05). Conclusion: The results indicate that protective effects of Artemisia extract against the thioacetamide-induced hepatotoxicity may be due to its ability to block the bioactivation of thioacetamide, primarily by inhibiting the activity of Cyp450 and free radicals. Artemisia possesses quercetin. Studies have demonstrated that quercetin inhibits lipid peroxidation and as an antioxidant can inhibit lipid peroxidation. PMID:25050286

  8. Distribution of tissue enzymes in three species of pinnipeds.

    PubMed

    Fauquier, Deborah A; Mazet, Jonna A K; Gulland, Frances M D; Spraker, Terry R; Christopher, Mary M

    2008-03-01

    In domestic animal medicine, changes in serum enzyme levels are routinely used as diagnostic tools to detect liver disease. Hepatic disease occurs in pinnipeds, but limited data are available on the tissue distribution of serum enzymes in marine mammals. The objectives of this study were to determine the tissue distribution of seven serum enzymes in three pinniped species. Enzymes evaluated were alanine aminotransferase (ALT), aspartate aminotransferase (AST), sorbitol dehydrogenase (SDH), lactate dehydrogenase (LDH), creatine kinase (CK), alkaline phosphatase (ALP), and gamma-glutamyl transferase (GGT) in tissues from California sea lions (Zalophus californianus) (n = 5), harbor seals (Phoca vitulina) (n = 5), and northern elephant seals (Mirounga angustirostris) (n = 5) that stranded and then died at a rehabilitation center. Samples were evaluated in duplicate from liver, skeletal muscle, cardiac muscle, kidney, adrenal, spleen, pancreas, lung, lymph node, and intestine. Patterns of tissue enzyme distribution were similar in all species, with SDH activity highest in liver and kidney, CK activity highest in skeletal and cardiac muscle, ALP activity highest in adrenal, and GGT activity highest in the kidney. Aspartate aminotransferase and LDH activities were less specific, with high activity in multiple tissues. Tissue ALT activity was high in the liver of all species, but was also high in cardiac muscle (California sea lions), skeletal muscle (harbor seals), and kidney (elephant seals). These results suggest that concurrent analysis of SDH, ALT, and CK would provide high specificity and sensitivity for the detection of hepatic lesions, and allow differentiation of liver from skeletal muscle lesions in pinniped species.

  9. Polymorphisms and features of cytomegalovirus UL144 and UL146 in congenitally infected neonates with hepatic involvement

    PubMed Central

    Ye, Sisi; Hu, Yingying; Li, Baoqing; Sun, Xiangwei; Mao, Chenchen; Xu, Jianfeng; Chen, Yiping; Zhang, Lifang; Xue, Xiangyang

    2017-01-01

    Human cytomegalovirus is a significant agent of hepatic involvement in neonates. In this study, we investigated the polymorphisms and features of the viral genes UL144 and UL146 as well as their significance to congenital hepatic involvement. In 79 neonates with congenital cytomegalovirus infection and hepatic involvement, full length UL144 and UL146 were successfully amplified in 73.42% and 60.76% of cases, respectively. Sequencing indicated that both genes were hypervariable. Notably, UL144 genotype B was highly associated with aspartate aminotransferase (P = 0.028) and lactate dehydrogenase (P = 0.046). Similarly, UL146 genotype G1 and G13 were significantly associated with CMV IgM (P = 0.026), CMV IgG (P = 0.034), alanine aminotransferase (P = 0.019), and aspartate aminotransferase (P = 0.032). In conclusion, dominant UL144 (genotype B) and UL146 (genotype G1 and G13) genotypes are associated with elevated levels of enzymes and CMV IgM and IgG of cytomegalovirus infection. PMID:28222150

  10. [Study of rat blood serum biochemical indicators of cardiotoxic action of novel antitumor 4-thiazolidinone derivatives and doxorubicin in complexes with polyethylene glycol-containing polymeric carrier in the rat blood serum].

    PubMed

    Kobylyns'ka, L I; Havryliuk, D Ia; Riabtseva, A O; Mitina, N Ie; Zaichenko, O S; Zimenkovskyĭ, B S; Stoĭka, R S

    2014-01-01

    The aim of this study was to measure the activity of enzymes which reflect cardiotoxic action in rats of novel synthetic 4-thiazolidone derivatives--3882, 3288 and 3833 that demonstrated antineoplastic effect in vitro towards 60 lines of human tumor cells tested in the framework of the program of screening new anticancer drugs at the National Cancer Institute (USA). Such action of these compounds was compared with the effect of well known anticancer agent doxorubicin and after conjugation of all above mentioned substances with new polyethylenglycol-containing polymeric comb-like carrier that was synthesized by the authors. Among the biochemical indicators of cardiotoxic action of anticancer agents, activity of the following enzymes in rat blood serum showed to be the most informative: creatine kinase, lactate dehydrogenase, aspartate aminotransferase, and alanine aminotransterase. Tenfold injection of doxorubicin in a dose of 5.5 mg/kg of weight caused rats' death, while 3882, 3288 and 3833 preparations had not such action. Application of the doxorubicin in combination with polymeric carrier prolonged the survival time to 20 days. Thus, the injection of anticancer agents in a complex with polymeric carrier provides a significant decrease in their cardiotoxicity that was confirmed by the corresponding changes in the activity of marker enzymes: creatine kinase, lactate dehydrogenase, aspartate aminotransferase and alanine aminotransferase in blood serum of treated rats.

  11. Clinicopathological features of serum TTV DNA-positive non-A-G liver diseases in Japan.

    PubMed

    Sioda, A; Moriyama, M; Matsumura, H; Kaneko, M; Tanaka, N; Arakawa, Y

    2001-10-01

    This study was undertaken to detect TTV DNA in serum samples from patients with non-A, non-B, non-C, non-E, and non-G (non-A-G) liver diseases and from blood donors, and to investigate the clinicopathological features of TTV infection including its prevalence and influence on liver disease. The study population consisted of 20 patients with non-A-G liver diseases (nine with chronic hepatitis (CH), six with liver cirrhosis (LC), and five with hepatocellular carcinoma (HCC), as well as 47 blood donors. Detection of TTV DNA was conducted with 200 &mgr;l of serum by the nested polymerase chain reaction. The detection rate of TTV DNA by subject category was CH 55.9; LC 66.7; HCC 60%; and blood donors 28%. Regarding blood biochemistry, TTV DNA-positive patients tended to show higher levels of aspartate aminotransferase and alanine aminotransferase, as well as lower levels of platelet counts. Long-term follow-up revealed that TTV DNA-positive patients exhibited characteristic, multiple peaks of alanine aminotransferase (ALT) levels. The histologic findings in the livers of TTV DNA-positive patients with CH consisted of moderate necro-inflammatory reactions. In conclusion, it is possible that the TTV genotype 1b infection caused liver injury.

  12. A contribution for the definition of serum chemistry values in captive adults Antillean manatees (Trichechus manatus manatus Linnaeus, 1758).

    PubMed

    Silva, F M O; Vergara-Parente, J E; Gomes, J K N; Teixeira, M N; Lima, R P

    2007-04-01

    Serum chemistry analyses represents a fundamental tool for the diagnosis and understanding of diseases in marine mammals. Although several studies are being conducted within the field of clinical pathology, haematological and serum chemistry data for Antillean manatees are deficient. The purpose of this study was to determine serum chemistry values for captive Antillean manatees within the CMA/Ibama facility in Brazil. Serum samples were obtained from five captive adult Antillean manatees fed with seagrass and analysed for aspartate aminotransferase, alanine aminotransferase, bilirubin, alkaline phosphatase, urea, creatinine, glucose, triglycerides, cholesterol, total protein, albumin, globulin, phosphate, chloride, calcium and uric acid. Blood chemistry parameters were determined using a semi-automatic analyzer. Maximum, minimum, mean and standard deviations were calculated for each serum chemistry parameter. Differences on the values of males and females were verified using an unpaired Student's t-test. All the parameters analysed were similar between sexes, with exception of AP, which was higher in females (191.43 +/- 31.86 U/l). Alanine aminotransferase and uric acid values for Trichechus manatus manatus are reported for the first time in this paper. This study is the first to report serum chemistry parameter values for long-term captive male and female Antillean manatees. Therefore, the lower values of albumin, phosphate, chloride, cholesterol and triglycerides obtained here highlight the importance of clinical pathology during health monitoring of captive marine mammals.

  13. Hepatotoxicity assessment of Mn-doped ZnS quantum dots after repeated administration in mice

    PubMed Central

    Yang, Yanjie; Lv, Shuang-Yu; Yu, Bianfei; Xu, Shuang; Shen, Jianmin; Zhao, Tong; Zhang, Haixia

    2015-01-01

    Doped ZnS quantum dots (QDs) have a longer dopant emission lifetime and potentially lower cytotoxicity compared to other doped QDs. The liver is the key organ for clearance and detoxification of xenobiotics by phagocytosis and metabolism. The present study was designed to synthesize and evaluate the hepatotoxicity of Mn-doped ZnS QDs and their polyethylene glycol-coated counterparts (1 mg/kg and 5 mg/kg) in mice. The results demonstrated that daily injection of Mn-doped ZnS QDs and polyethylene glycol-coated QDs via tail vein for 7 days did not influence body weight, relative liver weight, serum aminotransferases (alanine aminotransferase and aspartate aminotransferase), the levels of antioxidant enzymes (catalase, glutathione peroxidase, and superoxide dismutase), or malondialdehyde in the liver. Analysis of hepatocyte ultrastructure showed that Mn-doped ZnS QDs and polyethylene glycol-coated QDs mainly accumulated in mitochondria at 24 hours after repeated intravenous injection. No damage to cell nuclei or mitochondria was observed with either of the QDs. Our results indicate that Mn-doped ZnS QDs did not cause obvious damage to the liver. This study will assist in the development of Mn-doped ZnS QDs-based bioimaging and biomedical applications in the future. PMID:26396512

  14. Ethylene and 1-MCP regulate major volatile biosynthetic pathways in apple fruit.

    PubMed

    Yang, Xiaotang; Song, Jun; Du, Lina; Forney, Charles; Campbell-Palmer, Leslie; Fillmore, Sherry; Wismer, Paul; Zhang, Zhaoqi

    2016-03-01

    The effects of ethylene and 1-methylcyclopropene (1-MCP) on apple fruit volatile biosynthesis and gene expression were investigated. Statistical analysis identified 17 genes that changed significantly in response to ethylene and 1-MCP treatments. Genes encoding branched-chain amino acid aminotransferase (BCAT), aromatic amino acid aminotransferase (ArAT) and amino acid decarboxylases (AADC) were up-regulated during ripening and further enhanced by ethylene treatment. Genes related to fatty acid synthesis and metabolism, including acyl-carrier-proteins (ACPs), malonyl-CoA:ACP transacylase (MCAT), acyl-ACP-desaturase (ACPD), lipoxygenase (LOX), hydroperoxide lyase (HPL), alcohol dehydrogenase (ADH), pyruvate decarboxylase (PDC2), β-oxidation, acyl-CoA synthetase (ACS), enoyl-CoA hydratase (ECHD), acyl-CoA dehydrogenase (ACAD), and alcohol acyltransferases (AATs) also increased during ripening and in response to ethylene treatment. Allene oxide synthase (AOS), alcohol dehydrogenase 1 (ADH1), 3-ketoacyl-CoA thiolase and branched-chain amino acid aminotransferase 2 (BCAT2) decreased in ethylene-treated fruit. Treatment with 1-MCP and ethylene generally produced opposite effects on related genes, which provides evidence that regulation of these genes is ethylene dependent.

  15. Cardioprotective and Antioxidant Influence of Aqueous Extracts from Sesamum indicum Seeds on Oxidative Stress Induced by Cadmium in Wistar Rats

    PubMed Central

    Oyinloye, Babatunji Emmanuel; Ajiboye, Basiru Olaitan; Ojo, Oluwafemi Adeleke; Nwozo, Sarah Onyenibe; Kappo, Abidemi Paul

    2016-01-01

    Background: Oxidative stress has been implicated in the pathogenesis of several acute and chronic diseases of the heart as a result of indiscriminate exposure to cardiotoxic heavy metals. The study reported here was designed to evaluate the possible ameliorative effect of aqueous extracts from Sesamum indicum (SI) seeds on oxidative stress induced by cadmium (Cd) in Wistar rats. Materials and Methods: Daily administration of Cd (200 mg/L Cd as CdCl2) in the animals’ main drinking water for 21 days led to oxidative stress. Thereafter, the ameliorative effects were assessed by measuring biochemical parameters such as extent of lipid peroxidation (LPO), lipid profile, and enzymatic and nonenzymatic antioxidants, as well as serum aminotransferase activities. Results: Treatment with SI extract elicited notable reduction in serum total cholesterol, triglyceride, and low-density lipoprotein cholesterol levels as well as concomitant increase in high-density lipoprotein cholesterol. SI extract also reversed the elevations witnessed in serum aminotransferase activities, LPO level, and ameliorated enzymatic and nonenzymatic antioxidant status in the heart of Cd-exposed rats. Conclusion: Thus, SI appears to be an attractive candidate with potential for the novel treatment of cardiotoxicity and management of oxidative stress arising from Cd exposure. SUMMARY Cadmium (200 mg/L) exposure in drinking water caused pronounced oxidative stress and cardiac tissue damage in animal modelAqueous extract of Sesamum indicum (SI) seeds at a dose of 200 or 400 mg/kg body weight exhibited a significant reversal effect in all biochemical parameters measured such as extent of lipid peroxidation, lipid profile, and enzymatic and nonenzymatic antioxidants, as well as serum aminotransferase activitiesAqueous extract of SI seeds possess antioxidant and cardioprotective potential in a dose-dependent manner, thus conferring protection against oxidative stress induced by cadmium. Abbreviation used

  16. Maternal Factors Are Associated with the Expression of Placental Genes Involved in Amino Acid Metabolism and Transport

    PubMed Central

    Day, Pricilla E.; Ntani, Georgia; Crozier, Sarah R.; Mahon, Pam A.; Inskip, Hazel M.; Cooper, Cyrus; Harvey, Nicholas C.; Godfrey, Keith M.; Hanson, Mark A.; Lewis, Rohan M.; Cleal, Jane K.

    2015-01-01

    Introduction Maternal environment and lifestyle factors may modify placental function to match the mother’s capacity to support the demands of fetal growth. Much remains to be understood about maternal influences on placental metabolic and amino acid transporter gene expression. We investigated the influences of maternal lifestyle and body composition (e.g. fat and muscle content) on a selection of metabolic and amino acid transporter genes and their associations with fetal growth. Methods RNA was extracted from 102 term Southampton Women’s Survey placental samples. Expression of nine metabolic, seven exchange, eight accumulative and three facilitated transporter genes was analyzed using quantitative real-time PCR. Results Increased placental LAT2 (p = 0.01), y+LAT2 (p = 0.03), aspartate aminotransferase 2 (p = 0.02) and decreased aspartate aminotransferase 1 (p = 0.04) mRNA expression associated with pre-pregnancy maternal smoking. Placental mRNA expression of TAT1 (p = 0.01), ASCT1 (p = 0.03), mitochondrial branched chain aminotransferase (p = 0.02) and glutamine synthetase (p = 0.05) was positively associated with maternal strenuous exercise. Increased glutamine synthetase mRNA expression (r = 0.20, p = 0.05) associated with higher maternal diet quality (prudent dietary pattern) pre-pregnancy. Lower LAT4 (r = -0.25, p = 0.05) and aspartate aminotransferase 2 mRNA expression (r = -0.28, p = 0.01) associated with higher early pregnancy diet quality. Lower placental ASCT1 mRNA expression associated with measures of increased maternal fat mass, including pre-pregnancy BMI (r = -0.26, p = 0.01). Lower placental mRNA expression of alanine aminotransferase 2 associated with greater neonatal adiposity, for example neonatal subscapular skinfold thickness (r = -0.33, p = 0.001). Conclusion A number of maternal influences have been linked with outcomes in childhood, independently of neonatal size; our finding of associations between placental expression of transporter

  17. Electrolyte and plasma enzyme analyses during large-volume liposuction.

    PubMed

    Lipschitz, Avron H; Kenkel, Jeffrey M; Luby, Maureen; Sorokin, Evan; Rohrich, Rod J; Brown, Spencer A

    2004-09-01

    Substantial fluid shifts occur during liposuction as wetting solution is infiltrated subcutaneously and fat is evacuated, causing potential electrolyte imbalances. In the porcine model for large-volume liposuction, plasma aspartate aminotransferase and alanine transaminase levels were elevated following liposuction. These results raised concerns for possible mechanical injury and/or lidocaine-induced hepatocellular toxicity in a clinical setting. The first objective of this human model study was to explore the effect of the liposuction procedure on electrolyte balance. The second objective was to determine whether elevated plasma aminotransferase levels were observed subsequent to large-volume liposuction. Five female volunteers underwent three-stage, ultrasound-assisted liposuction. Blood samples were collected perioperatively. Plasma levels of sodium, potassium, venous carbon dioxide, blood urea nitrogen, chloride, and creatinine were determined. Liver function analyte levels were measured, including albumin, total protein, aspartate aminotransferase, and alanine transaminase, alkaline phosphatase, gamma-glutamyl transpeptidase, and total bilirubin. To further define intracellular enzyme release, creatine kinase levels were measured. Mild hyponatremia was evident postoperatively (134 to 136 mmol/liter) in four patients. Hypokalemia was evident intraoperatively in all subjects (mean +/- SEM; 3.3 +/- 0.16 mmol/liter; range, 3.0 to 3.4 mmol/liter). Hypoalbuminemia and hypoproteinemia were observed throughout the study (baseline: 2.9 +/- 0.2 g/dl; range, 2.6 to 3.5 g/dl), decreasing to 10 to 40 percent 24 hours postoperatively (2.0 +/- 0.2 g/dl; range, 1.7 to 2.1 g/dl). Aspartate aminotransferase, alanine transaminase, and creatine kinase levels were significantly elevated after the procedure (190 +/- 47.1 U/liter, 50 +/- 7.7 U/liter, and 11,219 +/- 2556.7 U/liter, respectively) (p < 0.01). Release of antidiuretic hormone and even mildly hypotonic intravenous fluid

  18. Hepatoprotective effects of erythropoietin on D-galactosamine/lipopolysaccharide-induced fulminant hepatic failure in mice.

    PubMed

    Yang, Xue-Fei; He, Yi; Li, Hai-Yuan; Liu, Xin; Chen, Huan; Liu, Jian-Bang; Ji, Wen-Jun; Wang, Bing; Chen, Li-Na

    2014-07-01

    Fulminant hepatic failure is a severe clinical syndrome associated with a high rate of patient mortality. Recent studies have shown that in addition to its hematopoietic effect, erythropoietin (EPO) has multiple protective effects and exhibits antiapoptotic, antioxidant and anti-inflammatory activities. The present study aimed to determine the hepatoprotective effect of EPO and to elucidate the underlying mechanisms using a D-galactosamine (D-GalN)/lipopolysaccharide (LPS)-induced model of acute liver injury. Experimental groups of mice were administered with various doses of EPO (1,000, 3,000 or 10,000 U/kg, intraperitoneal) once per day for 3 days, prior to injection with D-GalN (700 mg/kg)/LPS (10 µg/kg). Mice were sacrificed 8 h after treatment with D‑GalN/LPS. Liver function and histopathology, malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GSH‑Px) activities and EPO receptor (EPOR) and phosphatidylinositol 3-kinase (PI3K) mRNA expression were evaluated. D-GalN/LPS administration markedly induced liver injury, as evidenced by elevated levels of serum aminotransferases, as well as histopathological changes. Compared with the D-GalN/LPS group, pretreatment with EPO significantly decreased the levels of aspartate aminotransferase, alanine aminotransferase and MDA, and increased the activities of SOD and GSH-Px. Furthermore, the protective effects of EPO were paralleled by an upregulation in the mRNA expression of EPOR and PI3K. These data suggest that EPO can ameliorate D-GalN/LPS-induced acute liver injury by reducing oxidative stress and upregulating the mRNA expression of EPOR and PI3K.

  19. Iron Status and Metabolic Syndrome in Patients with Non-Alcoholic Fatty Liver Disease

    PubMed Central

    Ghamarchehreh, Mohammad Ebrahim; Jonaidi-Jafari, Nematollah; Bigdeli, Mohammad; Khedmat, Hossein; Saburi, Amin

    2016-01-01

    BACKGROUND A hypothesis has been presented about the role of serum iron, ferritin and transferrin saturation among patients with non-alcoholic fatty liver disease (NAFLD) and resistance to insulin (metabolic syndrome [MetS]), but there is much controversy. This study aimed at investigating the level of serum iron and demographic characteristics in patients with NAFLD with or without MetS. METHODS A case-control study was conducted on patients with elevated liver enzymes referring to Baqiyatallah clinic, Tehran, Iran during 2010-2011. After ruling out other causes of increased aminotransferases and approving the diagnosis of NAFLD, the patients were divided into two groups of with or without MetS. Then, the individuals’ demographic, sonographic, and laboratory characteristics were recorded. RESULTS This research included 299 patients suffering from NAFLD who were divided into MetS (n=143; 47.8%) and non-MetS (n=156; 52.2%) groups. The age, systolic and diastolic blood pressure, body mass index, waist/hip ratio, glucose tolerance test, serum insulin, C. peptide, triglyceride, and HB A1c were different between MetS and non-MetS groups (p<0.05). There was no significant difference in serum iron and ferritin levels between the two groups, however, a significant correlation was found between serum ferritin and alanine transaminase (p=0.005) and also aspartate aminotransferase (p=0.032). CONCLUSION Our findings did not show a significant relationship between iron, in free or storage form, and the presence of MetS among patients with NAFLD, but serum ferritin can correlate with hepatocytes injuries indicated by raised aminotransferases. Nevertheless, to clarify this relationship further molecular, genomic, and histopathological studies are required. PMID:26933479

  20. Natural and Synthetic Variants of the Tricarboxylic Acid Cycle in Cyanobacteria: Introduction of the GABA Shunt into Synechococcus sp. PCC 7002

    PubMed Central

    Zhang, Shuyi; Qian, Xiao; Chang, Shannon; Dismukes, G. C.; Bryant, Donald A.

    2016-01-01

    For nearly half a century, it was believed that cyanobacteria had an incomplete tricarboxylic acid (TCA) cycle, because 2-oxoglutarate dehydrogenase (2-OGDH) was missing. Recently, a bypass route via succinic semialdehyde (SSA), which utilizes 2-oxoglutarate decarboxylase (OgdA) and succinic semialdehyde dehydrogenase (SsaD) to convert 2-oxoglutarate (2-OG) into succinate, was identified, thus completing the TCA cycle in most cyanobacteria. In addition to the recently characterized glyoxylate shunt that occurs in a few of cyanobacteria, the existence of a third variant of the TCA cycle connecting these metabolites, the γ-aminobutyric acid (GABA) shunt, was considered to be ambiguous because the GABA aminotransferase is missing in many cyanobacteria. In this study we isolated and biochemically characterized the enzymes of the GABA shunt. We show that N-acetylornithine aminotransferase (ArgD) can function as a GABA aminotransferase and that, together with glutamate decarboxylase (GadA), it can complete a functional GABA shunt. To prove the connectivity between the OgdA/SsaD bypass and the GABA shunt, the gadA gene from Synechocystis sp. PCC 6803 was heterologously expressed in Synechococcus sp. PCC 7002, which naturally lacks this enzyme. Metabolite profiling of seven Synechococcus sp. PCC 7002 mutant strains related to these two routes to succinate were investigated and proved the functional connectivity. Metabolite profiling also indicated that, compared to the OgdA/SsaD shunt, the GABA shunt was less efficient in converting 2-OG to SSA in Synechococcus sp. PCC 7002. The metabolic profiling study of these two TCA cycle variants provides new insights into carbon metabolism as well as evolution of the TCA cycle in cyanobacteria. PMID:28018308

  1. Influence of estrogens on copper indicators: in vivo and in vitro studies.

    PubMed

    Arredondo, Miguel; Núñez, Héctor; López, Guadalupe; Pizarro, Fernando; Ayala, Mariana; Araya, Magdalena

    2010-06-01

    Classic copper indicators are not sensitive and specific for detecting excess copper exposure when this is higher than customary but not markedly elevated. Serum copper and ceruloplasmin (Cp) are the most commonly used indicators to assess nutritional status of copper. The objective of this paper was to study the influence of estrogens on these indicators and others used to assess early effects of excess copper exposure in humans and the expression of a set of copper related proteins in a hepatic cellular model. For the studies in humans, 107 healthy participants (18-50 years) were allocated as follows: group 1 (n = 39), women assessed on day 7 of their hormonal cycle; group 2 (n = 34), women assessed on day 21 of their hormonal cycle, and group 3 (n = 34, comparison group), healthy men. Participants received 8 mg Cu/day (as copper sulfate) during 6 months. Serum Cp and Cu, Cu-Zn-superoxide dismutase activity, liver function indicators [aspartate aminotransferase (AST), alanine aminotransferase (ALT), and gamma glutamyltransferase (GGT)], and serum Fe and Zn concentrations were measured monthly. In addition, the influence of estradiol on intracellular total copper content, hctr1, dmt1 and shbg mRNA abundance and hCTR1, and DMT1 expression was measured in HepG2 cells. Serum Cu, Fe, and Zn and liver aminotransferases but not Cu-Zn-superoxide dismutase varied depending on sex. Fe nutrition indicators, GGT, and ALT activities showed significant differences between the hormonal phases. Cellular experiments showed that estradiol increased cellular Cu concentration and hCTR1 and DMT1 mRNA expression and changed these proteins expression patterns. Estradiols significantly influence the responses to copper at the whole body and the cellular levels, suggesting that they help maintaining copper availability for metabolic needs.

  2. Effect of space flights on plasma hormone levels in man and in experimental animal

    NASA Astrophysics Data System (ADS)

    Macho, L.; Kvetňanský, R.; Vigaš, M.; Németh, S.; Popova, I.; Tigranian, R. A.; Noskov, V. B.; Serova, L.; Grigoriev, I. A.

    An important increase of plasma hormone levels like insulin, TSH and aldosterone was observed in human subjects after space flights, however in the changes of plasma content of ACTH, cortisol, adrenaline and noradrenaline the individual variations were observed in relation to number and duration of space flight. For evaluation of the effects of these changes in plasma hormone levels on metabolic processes also the experiments with small animals subjected to space flights on a board of biosatellite of Cosmos series were running. An elevation of plasma levels of corticosterone, adrenaline, noradrenaline and insulin was found in rats after the space flights of duration from 7 to 20 days. It was demonstrated, that the increase of corticosterone in plasma is followed by the activation of enzymes involved in the aminoacid metabolism in rat liver (tyrosine aminotransferase, tryptophanpyrolase, alanine aminotransferase and aspartate aminotransferase). After a short recovery period (2 to 6 days) the plasma corticosterone concentration and also the activity of liver enzymes returned to control levels. The exposition of animals to stress stimuli during this recovery period showed higher response of corticosterone levels in flight rats as compared to intact controls. The increase of plasma catecholamine levels was not followed by elevation of lipolysis in adipose tissue. This is due to lower response of adipose tissue to catecholamine because a decrease of the stimulation of lipolysis by noradrenaline was observed in animals after space flight. The increase of insulin was not followed by adequate decrease of glucose concentration suggesting a disturbances in glucose utilization similarly as in cosmonauts after a long-term space flight. These results showed that changes in plasma hormone levels, observed after space flight, affected the regulation of metabolic processes in tissues.

  3. Metabolic changes in the animals subjected to space flight

    NASA Astrophysics Data System (ADS)

    Macho, Ladislav; Németh, Štefan; Kvetňanský, Richard; Ficková, Maria; Tigranian, Ruben A.; Serova, Ljubov

    1982-07-01

    The activity of the enzymes involved in aminoacid metabolism (tyrosine aminotransferase, TAT, tryptophan pyrrolase TP, serine dehydratase, SD) with rapid response to glucocorticoids and enzymes requiring for activity increase repeated administration of corticosterone (alanine aminotransferase, ALT, aspartate aminotransferase, AST) in liver, the changes of lipolysis in adipose tissue and the plasma corticosterone levels were studied in rats subjected to space flight (F), in animals from synchron model experiments (SM, simulated conditions of space flight in laboratory) and in intact controls (C). The increase of plasma corticosterone concentration and of the activity of rapidly (TAT, TP, SD) and slowly activating enzymes (ALT, AST) was found in F group 6-10 hr after space flight (18.5 days on biosatellite COSMOS 1129). This suggested the presence of acute-stress (associated primarily with the landing) and chronic stress induced hypercorticosteronemia during the flight. After the short 6-day period of recovery the plasma corticosterone concentrations and the activities of liver enzymes returned to control levels. The exposition of animals to repeated immobilization stress showed higher response of corticosterone levels in flight rats as compared to intact controls. No changes in basal lipolysis were observed in flight rats in comparison to intact controls, however the stimulation of lipolysis by norepinephrine was lover in animals from F and SM groups. This lower response of lipolytic processes to norepinephrine was found in flight animals also after six days period of recovery. These results showed that there are important changes in the regulation of lipolytic processes in adipose tissue of rats after space flight and in the conditions of model experiments.

  4. Mitochondrial carnitine palmitoyl transferase-II inactivity aggravates lipid accumulation in rat hepatocarcinogenesis

    PubMed Central

    Gu, Juan-Juan; Yao, Min; Yang, Jie; Cai, Yin; Zheng, Wen-Jie; Wang, Li; Yao, Deng-Bing; Yao, Deng-Fu

    2017-01-01

    AIM To investigate the dynamic alteration of mitochondrial carnitine palmitoyl transferase II (CPT-II) expression during malignant transformation of rat hepatocytes. METHODS Sprague-Dawley male rats were fed with normal, high fat (HF), and HF containing 2-fluorenylacetamide (2-FAA) diet, respectively. According to the Hematoxylin and Eosin staining of livers, rats were divided into control, fatty liver, degeneration, precancerous, and cancerous groups. Liver lipids were dyed with Oil Red O, CPT-II alterations were analyzed by immunohistochemistry, and compared with CPT-II specific concentration (μg/mg protein). Levels of total cholesterol (Tch), triglyceride (TG), and amino-transferases [alanine aminotransferase (ALT), aspartate aminotransferase (AST)] were determined by the routine methods. RESULTS After intake of HF and/or HF+2-FAA diets, the rat livers showed mass lipid accumulation. The lipid level in the control group was significantly lower than that in other groups. The changes of serum TG and Tch levels were abnormally increasing, 2-3 times more than those in the controls (P < 0.05). During the rat liver morphological changes from normal to cancer development process with hepatocyte injury, serum AST and ALT levels were significantly higher (4-8 times, P < 0.05) than those in the control group. The specific concentration of CPT-II in liver tissues progressively decreased during hepatocyte malignant transformation, with the lowest CPT-II levels in the cancer group than in any of the other groups (P < 0.05). CONCLUSION Low CPT-II expression might lead to abnormal hepatic lipid accumulation, which should promote the malignant transformation of hepatocytes. PMID:28127199

  5. Immunochemical quantitation of 3-(cystein-S-yl)acetaminophen adducts in serum and liver proteins of acetaminophen-treated mice.

    PubMed

    Pumford, N R; Hinson, J A; Potter, D W; Rowland, K L; Benson, R W; Roberts, D W

    1989-01-01

    Using a recently developed enzyme-linked immunosorbent assay specific for 3-(cystein-S-yl)acetaminophen adducts we have quantitated the formation of these specific adducts in liver and serum protein of B6C3F1 male mice dosed with acetaminophen. Administration of acetaminophen at doses of 50, 100, 200, 300, 400 and 500 mg/kg to mice resulted in evidence of hepatotoxicity (increase in serum levels of alanine aminotransferase and aspartate aminotransferase) at 4 hr in the 300, 400 and 500 mg/kg treatment groups only. The formation of 3-(cystein-S-yl)acetaminophen adducts in liver protein was not observed in the groups receiving 50, 100 and 200 mg/kg doses, but was observed in the groups receiving doses above 300 mg/kg of acetaminophen. Greater levels of adduct formation were observed at the higher doses. 3-(Cystein-S-yl)acetaminophen protein adducts were also observed in serum of mice receiving hepatotoxic doses of acetaminophen. After a 400 mg/kg dose of acetaminophen, 3-(cystein-S-yl)acetaminophen adducts in the liver protein reached peak levels 2 hr after dosing. By 12 hr the levels decreased to approximately 10% of the peak level. In contrast, 3-(cystein-S-yl)acetaminophen adducts in serum protein were delayed, reaching a sustained peak 6 to 12 hr after dosing. The dose-response correlation between the appearance of serum aminotransferases and 3-(cystein-S-yl)acetaminophen adducts in serum protein and the temporal correlation between the decrease in 3-(cystein-S-yl)acetaminophen adducts in liver protein and the appearance of adducts in serum protein are consistent with a hepatic origin of the adducts detected in serum protein.(ABSTRACT TRUNCATED AT 250 WORDS)

  6. Conversion of Methionine to Thiols by Lactococci, Lactobacilli, and Brevibacteria†

    PubMed Central

    Dias, Benjamin; Weimer, Bart

    1998-01-01

    Methanethiol has been strongly associated with desirable Cheddar cheese flavor and can be formed from the degradation of methionine (Met) via a number of microbial enzymes. Methionine γ-lyase is thought to play a major role in the catabolism of Met and generation of methanethiol in several species of bacteria. Other enzymes that have been reported to be capable of producing methanethiol from Met in lactic acid bacteria include cystathionine β-lyase and cystathionine γ-lyase. The objective of this study was to determine the production, stability, and activities of the enzymes involved in methanethiol generation in bacteria associated with cheese making. Lactococci and lactobacilli were observed to contain high levels of enzymes that acted primarily on cystathionine. Enzyme activity was dependent on the concentration of sulfur amino acids in the growth medium. Met aminotransferase activity was detected in all of the lactic acid bacteria tested and α-ketoglutarate was used as the amino group acceptor. In Lactococcus lactis subsp. cremoris S2, Met aminotransferase was repressed with increasing concentrations of Met in the growth medium. While no Met aminotransferase activity was detected in Brevibacterium linens BL2, it possessed high levels of l-methionine γ-lyase that was induced by addition of Met to the growth medium. Met demethiolation activity at pH 5.2 with 4% NaCl was not detected in cell extracts but was detected in whole cells. These data suggest that Met degradation in Cheddar cheese will depend on the organism used in production, the amount of enzyme released during aging, and the amount of Met in the matrix. PMID:9726877

  7. Prevalence and characteristics of hypoxic hepatitis in the largest single-centre cohort of avian influenza A(H7N9) virus-infected patients with severe liver impairment in the intensive care unit

    PubMed Central

    Zhang, YiMin; Liu, JiMin; Yu, Liang; Zhou, Ning; Ding, Wei; Zheng, ShuFa; Shi, Ding; Li, LanJuan

    2016-01-01

    Avian influenza A(H7N9) virus (A(H7N9)) emerged in February 2013. Liver impairment of unknown cause is present in 29% of patients with A(H7N9) infection, some of whom experience severe liver injury. Hypoxic hepatitis (HH) is a type of acute severe liver injury characterized by an abrupt, massive increase in serum aminotransferases resulting from anoxic centrilobular necrosis of liver cells. In the intensive care unit (ICU), the prevalence of HH is ∼1%–2%. Here, we report a 1.8% (2/112) incidence of HH in the largest single-centre cohort of ICU patients with A(H7N9) infection. Both HH patients presented with multiple organ failure (MOF) involving respiratory, cardiac, circulatory and renal failure and had a history of chronic heart disease. On admission, severe liver impairment was found. Peak alanine aminotransferase (ALT) and aspartate aminotransferase (AST) values were 937 and 1281 U/L, and 3117 and 3029 U/L, respectively, in the two patients. Unfortunately, both patients died due to deterioration of MOF. A post-mortem biopsy in case 1 confirmed the presence of centrilobular necrosis of the liver, and real-time reverse transcription polymerase chain reaction of A(H7N9)-specific genes was negative, which excluded A(H7N9)-related hepatitis. The incidence of HH in A(H7N9) patients is similar to that in ICU patients with other aetiologies. It seems that patients with A(H7N9) infection and a history of chronic heart disease with a low left ventricular ejection fraction on admission are susceptible to HH, which presents as a marked elevation in ALT at the time of admission. PMID:26733380

  8. Cord Blood Adipokines and Lipids and Adolescent Nonalcoholic Fatty Liver Disease

    PubMed Central

    Simpson, Joy; Smith, Andrew D.; Fraser, Abigail; Sattar, Naveed; Callaway, Mark; Lindsay, Robert S.; Lawlor, Debbie A.

    2016-01-01

    Context: Maternal adiposity in pregnancy is associated with offspring adiposity and metabolic dysfunction postnatally, including greater risk of nonalcoholic fatty liver disease (NAFLD). Recent genetic analyses suggest a causal effect of greater maternal body mass index on offspring birth weight and ponderal index, but the relative roles of the environment in utero or later in life remains unclear. Objective: We sought to determine whether markers of infant adiposity (birth weight, umbilical cord blood leptin, adiponectin, and lipids) were associated with markers of NAFLD in adolescence. Design, Setting, and Participants: This was a UK prospective birth cohort with 17 years of follow-up with liver function tests (aspartate aminotransferase, alanine aminotransferase, gamma-glutamyltransferase) (n = 1037 participants), and ultrasound scan assessed liver fat, volume, and sheer velocity at age 17 (n = 541 participants). Missing covariate data were imputed. Main Outcomes: Ultrasound and biochemical measures of NAFLD were measured. Results: Birth weight, cord blood leptin, and adiponectin were not associated with a diagnosis of NAFLD. In adjusted analyses, 2 of 42 associations attained conventional 5% levels of significance. Birth weight was positively associated with liver volume (1.0% greater per 100 g [95% confidence interval 0.5%–2.0%]). Cord high-density lipoprotein-cholesterol was positively associated with alanine aminotransferase (11.6% higher per 1 mmol/L [95% confidence interval 0.3, 23.4]); however, this association was primarily mediated via offspring adiposity. Conclusions: In this extensive analysis, we found little evidence measurements of infant fat mass and birth size were related to adolescent markers of NAFLD. The association between birth weight and adolescent liver volume may indicate the contribution of greater organ size to birth weight and tracking of organ size. PMID:27648968

  9. Biochemical, anthropometric and body composition indicators as predictors of hepatic steatosis in obese adolescents

    PubMed Central

    Gobato, Amanda Oliva; Vasques, Ana Carolina J.; Yamada, Roberto Massao; Zambon, Mariana Porto; Barros-Filho, Antonio de Azevedo; Hessel, Gabriel

    2014-01-01

    OBJECTIVE: To describe the prevalence of hepatic steatosis and to assess the performance of biochemical, anthropometric and body composition indicators for hepatic steatosis in obese teenagers. METHODS: Cross-sectional study including 79 adolecents aged from ten to 18 years old. Hepatic steatosis was diagnosed by abdominal ultrasound in case of moderate or intense hepatorenal contrast and/or a difference in the histogram ≥7 on the right kidney cortex. The insulin resistance was determined by the Homeostasis Model Assessment-Insulin Resistance (HOMA-IR) index for values >3.16. Anthropometric and body composition indicators consisted of body mass index, body fat percentage, abdominal circumference and subcutaneous fat. Fasting glycemia and insulin, lipid profile and hepatic enzymes, such as aspartate aminotransferase, alanine aminotransferase, gamma-glutamyltransferase and alkaline phosphatase, were also evaluated. In order to assess the performance of these indicators in the diagnosis of hepatic steatosis in teenagers, a ROC curve analysis was applied. RESULTS: Hepatic steatosis was found in 20% of the patients and insulin resistance, in 29%. Gamma-glutamyltransferase and HOMA-IR were good indicators for predicting hepatic steatosis, with a cutoff of 1.06 times above the reference value for gamma-glutamyltransferase and 3.28 times for the HOMA-IR. The anthropometric indicators, the body fat percentage, the lipid profile, the glycemia and the aspartate aminotransferase did not present significant associations. CONCLUSIONS: Patients with high gamma-glutamyltransferase level and/or HOMA-IR should be submitted to abdominal ultrasound examination due to the increased chance of having hepatic steatosis. PMID:25119755

  10. Mercaptopurine/Methotrexate maintenance therapy of childhood acute lymphoblastic leukemia: clinical facts and fiction.

    PubMed

    Schmiegelow, Kjeld; Nielsen, Stine N; Frandsen, Thomas L; Nersting, Jacob

    2014-10-01

    The antileukemic mechanisms of 6-mercaptopurine (6MP) and methotrexate (MTX) maintenance therapy are poorly understood, but the benefits of several years of myelosuppressive maintenance therapy for acute lymphoblastic leukemia are well proven. Currently, there is no international consensus on drug dosing. Because of significant interindividual and intraindividual variations in drug disposition and pharmacodynamics, vigorous dose adjustments are needed to obtain a target degree of myelosuppression. As the normal white blood cell counts vary by patients' ages and ethnicity, and also within age groups, identical white blood cell levels for 2 patients may not reflect the same treatment intensity. Measurements of intracellular levels of cytotoxic metabolites of 6MP and MTX can identify nonadherent patients, but therapeutic target levels remains to be established. A rise in serum aminotransferase levels during maintenance therapy is common and often related to high levels of methylated 6MP metabolites. However, except for episodes of hypoglycemia, serious liver dysfunction is rare, the risk of permanent liver damage is low, and aminotransferase levels usually normalize within a few weeks after discontinuation of therapy. 6MP and MTX dose increments should lead to either leukopenia or a rise in aminotransferases, and if neither is experienced, poor treatment adherence should be considered. The many genetic polymorphisms that determine 6MP and MTX disposition, efficacy, and toxicity have precluded implementation of pharmacogenomics into treatment, the sole exception being dramatic 6MP dose reductions in patients who are homozygous deficient for thiopurine methyltransferase, the enzyme that methylates 6MP and several of its metabolites. In conclusion, maintenance therapy is as important as the more intensive and toxic earlier treatment phases, and often more challenging. Ongoing research address the applicability of drug metabolite measurements for dose adjustments

  11. Protective effects of medical ozone combined with traditional Chinese medicine against chemically-induced hepatic injury in dogs

    PubMed Central

    Li, Li-Jie; Yang, Yun-Gao; Zhang, Zhi-Ling; Nie, Sui-Feng; Li, Ze; Li, Feng; Hua, He-Yu; Hu, Yan-Jun; Zhang, Hong-Shuan; Guo, Ya-Bing

    2007-01-01

    AIM: To investigate the protective effect of medical ozone (O3) combined with Traditional Chinese Medicine (TCM) Yigan Fuzheng Paidu Capsules (YC) against carbon tetrachloride (CCl4)-induced hepatic injury in dogs. METHODS: Thirty healthy dogs were divided randomly into five groups (n = 6 in each group), namely control, oleanolic acid tablet (OAT), O3, YC and O3 + YC, given either no particular pre-treatment, oral OAT, medical ozone rectal insulfflation every other day, oral YC, or oral YC plus medical ozone rectal insulfflation every other day, respectively, for 30 consecutive days. After pre-treatment, acute hepatic injury was induced in all dogs with a single-dose intraperitoneal injection of CCl4. General condition and survival time were recorded. The biochemical and hematological indexes of alanine aminotransferase (ALT), aspartate aminotransferase/alanine aminotransferase (AST/ALT), serum total bilirubin (TBIL), prothrombin time (PT), blood ammonia (AMMO), and blood urea nitrogen (BUN) were measured after CCl4 injection. Hepatic pathological changes were also observed. RESULTS: Compared to the other four groups, the changes of group O3 + YC dogs’ general conditions (motoricity, mental state, eating, urination and defecation) could be better controlled. In group O3 + YC the survival rates were higher (P < 0.05 vs group control). AST/ALT values were kept within a normal level in group O3 + YC. Hepatic histopathology showed that hepatic injury in group O3 + YC was less serious than those in the other four groups. CONCLUSION: Medical ozone combined with TCM YC could exert a protective effect on acute liver injury induced by CCl4. PMID:18023088

  12. Prevalence and characteristics of hypoxic hepatitis in the largest single-centre cohort of avian influenza A(H7N9) virus-infected patients with severe liver impairment in the intensive care unit.

    PubMed

    Zhang, YiMin; Liu, JiMin; Yu, Liang; Zhou, Ning; Ding, Wei; Zheng, ShuFa; Shi, Ding; Li, LanJuan

    2016-01-06

    Avian influenza A(H7N9) virus (A(H7N9)) emerged in February 2013. Liver impairment of unknown cause is present in 29% of patients with A(H7N9) infection, some of whom experience severe liver injury. Hypoxic hepatitis (HH) is a type of acute severe liver injury characterized by an abrupt, massive increase in serum aminotransferases resulting from anoxic centrilobular necrosis of liver cells. In the intensive care unit (ICU), the prevalence of HH is ∼1%-2%. Here, we report a 1.8% (2/112) incidence of HH in the largest single-centre cohort of ICU patients with A(H7N9) infection. Both HH patients presented with multiple organ failure (MOF) involving respiratory, cardiac, circulatory and renal failure and had a history of chronic heart disease. On admission, severe liver impairment was found. Peak alanine aminotransferase (ALT) and aspartate aminotransferase (AST) values were 937 and 1281 U/L, and 3117 and 3029 U/L, respectively, in the two patients. Unfortunately, both patients died due to deterioration of MOF. A post-mortem biopsy in case 1 confirmed the presence of centrilobular necrosis of the liver, and real-time reverse transcription polymerase chain reaction of A(H7N9)-specific genes was negative, which excluded A(H7N9)-related hepatitis. The incidence of HH in A(H7N9) patients is similar to that in ICU patients with other aetiologies. It seems that patients with A(H7N9) infection and a history of chronic heart disease with a low left ventricular ejection fraction on admission are susceptible to HH, which presents as a marked elevation in ALT at the time of admission.

  13. Analysis of weaning-induced stress in Saanen goat kids.

    PubMed

    Magistrelli, D; Aufy, A A; Pinotti, L; Rosi, F

    2013-08-01

    In young ruminants' life, weaning often coincides with a period of growth stasis and poor welfare. The present study aimed at evaluating the effect of coping with the new diet on behavioural and haematological stress indicators in goat kids subjected to a commonly adopted weaning practice. Immediately after birth, male Saanen goat kids were divided into two groups: MILK and WMIX. All were fed colostrum for the first 3 days and then goat milk to the age of 29 days. After that, MILK kids continued to receive milk, while the WMIX kids underwent weaning and were completely weaned by day 48. Animal behaviour was recorded daily. From day 23-50, blood samples were taken weekly and analysed for indicators of stress and immune function. No abnormal behaviour, such as injurious behaviours or stereotypies, was observed in either of the experimental groups throughout the experimental period. During the last week, fasting plasma cortisol level was significantly lower, whereas plasma activity of both alanine aminotransferase (ALT) and aspartate aminotransferase (AST) was significantly higher in WMIX kids, in relation to the MILK ones. Anyway, data were within the normal physiological range and no difference was observed neither in plasma haptoglobin, ceruloplasmin, albumin and antithrombin III, nor in plasma immunoglobulin A and G, at any time, signalling no stressful condition. Therefore, differences observed in cortisol, ALT and AST could be the consequence of the metabolic changes that occur during the transition from pre-ruminant to ruminant state. The gradual weaning at 48 days of age did not result in any stressful condition and had no negative effect on weight gain. Results suggest that parameters commonly adopted to provide information on animal stress, such as cortisol and aminotransferase activity, can vary in relation to the physiological status of the animals and may bias stress assessment.

  14. Cholestatic jaundice, acute kidney injury and acute pancreatitis secondary to the recreational use of methandrostenolone: a case report

    PubMed Central

    2011-01-01

    Introduction Over the last few years the use of anabolic steroids has become increasingly common amongst amateur athletes and for aesthetic purposes. As a result, the adverse events related to their use are being seen more frequently. Methandrostenolone is an anabolic steroid which is widely available and has been used for both performance enhancement and aesthetic purposes. This drug has also been reported to cause cholestasis of the intra-hepatic bile ducts resulting in elevated aminotransferases, hyperbilirubinemia and clinical jaundice. However, to the best of our knowledge this agent has not been previously reported to cause pancreatitis or acute kidney injury. Case presentation In this paper, we report the case of a 50-year-old man of Indian descent who presented with a six week history of diffuse abdominal pain, anorexia and weight loss following an eight week cycle of methandrostenolone use. At initial presentation, his lipase level was 785 U/L, bilirubin was 922 μmol/L and creatinine was 200 U/L while his aspartate aminotransferase and alanine aminotransferase levels were only mildly elevated at 61 U/L and 56 U/L respectively. His lipase peaked on day nine at >3000 U/L whilst his creatinine level was 299 U/L. Imaging was consistent with acute pancreatitis while a liver biopsy was consistent with intra-hepatic cholestasis and a kidney biopsy revealed evidence of acute tubular necrosis. Conclusion Both acute pancreatitis and acute kidney injury have rarely been reported with anabolic steroid use and they have not been previously reported to occur in the same patient. This case demonstrates some potentially new and serious adverse consequences occurring with the use of anabolic steroids, of which physicians need to be aware. PMID:21470406

  15. A Small-Molecule Screen Identifies l-Kynurenine as a Competitive Inhibitor of TAA1/TAR Activity in Ethylene-Directed Auxin Biosynthesis and Root Growth in Arabidopsis[C][W

    PubMed Central

    He, Wenrong; Brumos, Javier; Li, Hongjiang; Ji, Yusi; Ke, Meng; Gong, Xinqi; Zeng, Qinglong; Li, Wenyang; Zhang, Xinyan; An, Fengying; Wen, Xing; Li, Pengpeng; Chu, Jinfang; Sun, Xiaohong; Yan, Cunyu; Yan, Nieng; Xie, De-Yu; Raikhel, Natasha; Yang, Zhenbiao; Stepanova, Anna N.; Alonso, Jose M.; Guo, Hongwei

    2011-01-01

    The interactions between phytohormones are crucial for plants to adapt to complex environmental changes. One example is the ethylene-regulated local auxin biosynthesis in roots, which partly contributes to ethylene-directed root development and gravitropism. Using a chemical biology approach, we identified a small molecule, l-kynurenine (Kyn), which effectively inhibited ethylene responses in Arabidopsis thaliana root tissues. Kyn application repressed nuclear accumulation of the ETHYLENE INSENSITIVE3 (EIN3) transcription factor. Moreover, Kyn application decreased ethylene-induced auxin biosynthesis in roots, and TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS1/TRYPTOPHAN AMINOTRANSFERASE RELATEDs (TAA1/TARs), the key enzymes in the indole-3-pyruvic acid pathway of auxin biosynthesis, were identified as the molecular targets of Kyn. Further biochemical and phenotypic analyses revealed that Kyn, being an alternate substrate, competitively inhibits TAA1/TAR activity, and Kyn treatment mimicked the loss of TAA1/TAR functions. Molecular modeling and sequence alignments suggested that Kyn effectively and selectively binds to the substrate pocket of TAA1/TAR proteins but not those of other families of aminotransferases. To elucidate the destabilizing effect of Kyn on EIN3, we further found that auxin enhanced EIN3 nuclear accumulation in an EIN3 BINDING F-BOX PROTEIN1 (EBF1)/EBF2-dependent manner, suggesting the existence of a positive feedback loop between auxin biosynthesis and ethylene signaling. Thus, our study not only reveals a new level of interactions between ethylene and auxin pathways but also offers an efficient method to explore and exploit TAA1/TAR-dependent auxin biosynthesis. PMID:22108404

  16. Comparison between cryoablation and irreversible electroporation of rabbit livers at a location close to the gallbladder

    PubMed Central

    Qin, Zilin; Zhou, Liang; Fang, Gang; Chen, Jibing; Li, Jialiang; Niu, Lizhi; Liang, Bing; Xu, Kecheng

    2017-01-01

    Abstract Background The ablation of liver tumors located close to the gallbladder is likely to lead to complications. The aim of this article is to compare the safety and efficacy of irreversible electroporation (IRE) and cryoablation in rabbit livers at a location close to the gallbladder. Materials and methods We performed cryoablation (n = 12) and IRE (n = 12) of the area of the liver close to the gallbladder in 24 New Zealand white rabbits in order to ensure gallbladder damage. Serum aminotransferase and serum bilirubin levels were measured before and after the ablation. Histopathological examination of the ablation zones in the liver and gallbladder was performed on the 7th day after the ablation. Result Seven days after the ablation, all 24 animals were alive. Gallbladder perforation did not occur in the IRE group; only mucosal epithelial necrosis and serous layer edema were found in this group. Gallbladder perforation occurred in four rabbits in the cryoablation group. Serum aminotransferase and serum bilirubin levels obviously increased in both groups by Day 3 and decreased gradually thereafter. The elevation in aminotransferase and bilirubin levels was greater in the cryoablation group than the IRE group. Pathological examination revealed complete necrosis of the liver parenchyma from the ablation center to the gallbladder in both groups, but bile duct and granulation tissue hyperplasia were observed in only the IRE group. Full-thickness gallbladder-wall necrosis was seen in the cryoablation group. Conclusions For ablation of the liver area near the gallbladder, IRE is superior to cryoablation, both in terms of safety (no gallbladder perforation in the IRE group) and efficacy (complete necrosis and rapid recovery in the IRE group). PMID:28265231

  17. Prediction and Diagnosis of Non-Alcoholic Fatty Liver Disease (NAFLD) and Identification of Its Associated Factors Using the Classification Tree Method

    PubMed Central

    Birjandi, Mehdi; Ayatollahi, Seyyed Mohammad Taghi; Pourahmad, Saeedeh; Safarpour, Ali Reza

    2016-01-01

    Background Non-alcoholic fatty liver disease (NAFLD) is the most common form of liver disease in many parts of the world. Objectives The aim of the present study was to identify the most important factors influencing NAFLD using a classification tree (CT) to predict the probability of NAFLD. Patients and Methods This cross-sectional study was conducted in Kavar, a town in the south of Fars province, Iran. A total of 1,600 individuals were selected for the study via the stratified method and multiple-stage cluster random sampling. A total of 30 demographic and clinical variables were measured for each individual. Participants were divided into two datasets: testing and training. We used the training dataset (1,120 individuals) to build the CT and the testing dataset (480 individuals) to assess the CT. The CT was also used to estimate class and to predict fatty liver occurrence. Results NAFLD was diagnosed in 22% of the individuals in the sample. Our findings revealed that the following variables, based on univariate analysis, had a significant association with NAFLD: marital status, history of hepatitis B vaccine, history of surgery, body mass index (BMI), waist-hip ratio (WHR), systolic blood pressure (SBP), diastolic blood pressure (DBP), high-density lipoprotein (HDL), triglycerides (TG), alanine aminotransferase (ALT), cholesterol (CHO0, aspartate aminotransferase (AST), glucose (GLU), albumin (AL), and age (P < 0.05). The main affecting variables for predicting NAFLD based on the CT and in order of importance were as follows: BMI, WHR, triglycerides, glucose, SBP, and alanine aminotransferase. The goodness of fit model based on the training and testing datasets were as follows: prediction accuracy (80%, 75%), sensitivity (74%, 73%), specificity (83%, 77%), and the area under the receiver operating characteristic (ROC) curve (78%, 75%), respectively. Conclusions The CT is a suitable and easy-to-interpret approach for decision-making and predicting NAFLD. PMID

  18. Lead and PCB's in canvasback ducks: Relationship between enzyme levels and residues in blood

    USGS Pu