Alanine Glyoxylate Aminotransferase

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CJ Danpure - One of the best experts on this subject based on the ideXlab platform.

  • Peroxisomal import of human Alanine : Glyoxylate Aminotransferase requires ancillary targeting information remote from its C terminus
    J BIOL CHEM, 2005
    Co-Authors: CJ Danpure
    Abstract:

    Although human Alanine: Glyoxylate Aminotransferase (AGT) is imported into peroxisomes by a Pex5p-dependent pathway, the properties of its C-terminal tripeptide (KKL) are unlike those of any other type 1 peroxisomal targeting sequence (PTS1). We have previously suggested that AGT might possess ancillary targeting information that enables its unusual PTS1 to work. In this study, we have attempted to locate this information and to determine whether or not it is a characteristic of all vertebrate AGTs. Using the two-hybrid system, we show that human AGT interacts with human Pex5p in mammalian cells, but not yeast cells. Using (immuno) fluorescence microscopic analysis of the distribution of various constructs expressed in COS cells, we show the following. 1) The putative ancillary peroxisomal targeting information (PTS1A) in human AGT is located entirely within the smaller C-terminal structural domain of 110 amino acids, with the sequence between Val-324 and Ile-345 being the most likely candidate region. 2) The PTS1A is present in all mammalian AGTs studied (human, rat, guinea pig, rabbit, and cat), but not amphibian AGT (Xenopus). 3) The PTS1A is necessary for peroxisomal import of human, rabbit, and cat AGTs, but not rat and guinea pig AGTs. We speculate that the internal PTS1A of human AGT works in concert with the C-terminal PTS1 by interacting with Pex5p indirectly with the aid of a yet-to-be-identified mammal-specific adaptor molecule. This interaction might reshape the tetratricopeptide repeat domain allosterically, enabling it to accept KKL as a functional PTS1.

  • Molecular evolution of Alanine : Glyoxylate Aminotransferase intracellular targeting
    2000
    Co-Authors: Jd Holbrook, CJ Danpure
    Abstract:

    Under the influence of dietary selection pressure, the subcellular distribution of the pyridoxal-phosphate-dependent enzyme Alanine:Glyoxylate Aminotransferase (AGT) has changed on numerous occasions throughout the evolution of mammals. Thus, AGT is mitochondrial in carnivores, peroxisomal in herbivores, and both mitochondrial and peroxisomal in omnivores. The variable distribution of AGT results from the variable use of two alternative transcription and translation start sites, such that an N-terminal mitochondrial targeting sequence is either included or excluded from the open reading frame.

  • Mammalian Alanine/Glyoxylate Aminotransferase 1 is imported into peroxisomes via the PTS1 translocation pathway. Increased degeneracy and context specificity of the mammalian PTS1 motif and implications for the peroxisome-to-mitochondrion mistargetin
    The Journal of cell biology, 1995
    Co-Authors: Alison M. Motley, P R Jennings, M. J. Lumb, P. B. Oatey, P. A. De Zoysa, Ronald J.a. Wanders, Henk F. Tabak, CJ Danpure
    Abstract:

    Alanine/Glyoxylate Aminotransferase 1 (AGT) is peroxisomal in most normal humans, but in some patients with the hereditary disease primary hyperoxaluria type 1 (PH1), AGT is mislocalized to the mitochondria. In an attempt to identify the sequences in AGT that mediate its targeting to peroxisomes, and to determine the mechanism by which AGT is mistargeted in PH1, we have studied the intracellular compartmentalization of various normal and mutant AGT polypeptides in normal human fibroblasts and cell lines with selective deficiencies of peroxisomal protein import, using immunofluorescence microscopy after intranuclear microinjection of AGT expression plasmids. The results show that AGT is imported into peroxisomes via the peroxisomal targeting sequence type 1 (PTS1) translocation pathway. Although the COOH-terminal KKL of human AGT was shown to be necessary for its peroxisomal import, this tripeptide was unable to direct the peroxisomal import of the bona fide peroxisomal protein firefly luciferase or the reporter protein bacterial chloramphenicol acetyltransferase. An ill-defined region immediately upstream of the COOH-terminal KKL was also found to be necessary for the peroxisomal import of AGT, but again this region was found to be insufficient to direct the peroxisomal import of chloramphenicol acetyltransferase. Substitution of the COOH-terminal KKL of human AGT by the COOH-terminal tripeptides found in the AGTs of other mammalian species (SQL, NKL), the prototypical PTS1 (SKL), or the glycosomal PTS1 (SSL) also allowed peroxisomal targeting, showing that the allowable PTS1 motif in AGT is considerably more degenerate than, or at least very different from, that acceptable in luciferase. AGT possessing the two amino acid substitutions responsible for its mistargeting in PH1 (i.e., Pro11-->Leu and Gly170-->Arg) was targeted mainly to the mitochondria. However, AGTs possessing each amino acid substitution on its own were targeted normally to the peroxisomes. This suggests that Gly170-->Arg-mediated increased functional efficiency of the otherwise weak mitochondrial targeting sequence (generated by the Pro11-->Leu polymorphism) is not due to interference with the peroxisomal targeting or import of AGT.

  • mammalian Alanine Glyoxylate Aminotransferase 1 is imported into peroxisomes via the pts1 translocation pathway increased degeneracy and context specificity of the mammalian pts1 motif and implications for the peroxisome to mitochondrion mistargeting
    Journal of Cell Biology, 1995
    Co-Authors: Alison M. Motley, P R Jennings, M. J. Lumb, P. B. Oatey, P. A. De Zoysa, Ronald J.a. Wanders, Henk F. Tabak, CJ Danpure
    Abstract:

    Alanine/Glyoxylate Aminotransferase 1 (AGT) is peroxisomal in most normal humans, but in some patients with the hereditary disease primary hyperoxaluria type 1 (PH1), AGT is mislocalized to the mitochondria. In an attempt to identify the sequences in AGT that mediate its targeting to peroxisomes, and to determine the mechanism by which AGT is mistargeted in PH1, we have studied the intracellular compartmentalization of various normal and mutant AGT polypeptides in normal human fibroblasts and cell lines with selective deficiencies of peroxisomal protein import, using immunofluorescence microscopy after intranuclear microinjection of AGT expression plasmids. The results show that AGT is imported into peroxisomes via the peroxisomal targeting sequence type 1 (PTS1) translocation pathway. Although the COOH-terminal KKL of human AGT was shown to be necessary for its peroxisomal import, this tripeptide was unable to direct the peroxisomal import of the bona fide peroxisomal protein firefly luciferase or the reporter protein bacterial chloramphenicol acetyltransferase. An ill-defined region immediately upstream of the COOH-terminal KKL was also found to be necessary for the peroxisomal import of AGT, but again this region was found to be insufficient to direct the peroxisomal import of chloramphenicol acetyltransferase. Substitution of the COOH-terminal KKL of human AGT by the COOH-terminal tripeptides found in the AGTs of other mammalian species (SQL, NKL), the prototypical PTS1 (SKL), or the glycosomal PTS1 (SSL) also allowed peroxisomal targeting, showing that the allowable PTS1 motif in AGT is considerably more degenerate than, or at least very different from, that acceptable in luciferase. AGT possessing the two amino acid substitutions responsible for its mistargeting in PH1 (i.e., Pro11-->Leu and Gly170-->Arg) was targeted mainly to the mitochondria. However, AGTs possessing each amino acid substitution on its own were targeted normally to the peroxisomes. This suggests that Gly170-->Arg-mediated increased functional efficiency of the otherwise weak mitochondrial targeting sequence (generated by the Pro11-->Leu polymorphism) is not due to interference with the peroxisomal targeting or import of AGT.

  • PRIMARY HYPEROXALURIA TYPE-1 AND PEROXISOME-TO-MITOCHONDRION MISTARGETING OF Alanine - Glyoxylate Aminotransferase
    Biochimie, 1993
    Co-Authors: CJ Danpure
    Abstract:

    Abstract Under the influence of dietary selection pressure, the intracellular compartmentalization of Alanine:Glyoxylate amino-transferase (AGT) has changed on many occasions during the evolution of mammals. In some mammals, AGT is peroxisomal in others it is mainly mitochondrial while in yet others it is more-or-less equally divided between both organelles. Although in normal human liver AGT is usually found exclusively within the peroxisomes, in some individuals a small proportion (≈5%) is found also in the mitochondria. This apparently trivial intracellular redistribution of AGT is caused by the presence of a Pro11Leu polymorphism which allows the N-terminus of AGT to fold into a conformation (ie a positively-charged amphiphilic α-helix) which functions as a mitochondrial targeting sequence. In one third of patients with the autosomal recessive disease primary hyperoxaluria type 1, there is a further redistribution of AGT so that the great majority (≈90%) is located in the mitochondria and only a small minority (10%) in the peroxisomes. AGT cannot fulfil its proper metabolic role in human liver (ie Glyoxylate detoxification) when located in the mitochondria. This erroneous compartmentalization is due to the presence of a Gly170Arg mutation superimposed upon the Pro11Leu polymorphism. The Gly170Arg mutation appears to have no direct effect on mitochondrial targeting and is predicted tlo enhance mitochondrial important of AGT by interfering with its peroxisomal targeting and/or import. The mitochondrial targeting sequence generated by the Pro11Leu polymorphism is not homologous to that found in the AGT of other mammals which localise AGT within the mitochondria normally. The identity of the peroxisomal targeting sequence in AGT is unkown, but the Gly170Arg mutation is found in a highly conserved region of the protein which might be involved in some aspect of the peroxisomal import pathway for AGT.

Yangdou Wei - One of the best experts on this subject based on the ideXlab platform.

  • Alanine: Glyoxylate Aminotransferase 1 is required for mobilization and utilization of triglycerides during infection process of the rice blast pathogen, Magnaporthe oryzae.
    Plant signaling & behavior, 2012
    Co-Authors: Vijai Bhadauria, Sabine Banniza, Albert Vandenberg, Gopalan Selvaraj, Yangdou Wei
    Abstract:

    The rice blast pathogen, Magnaporthe oryzae has been widely used as a model pathogen to study plant infection-related fungal morphogenesis, such as penetration via appressorium and plant-microbe interactions at the molecular level. Previously, we identified a gene encoding peroxisomal Alanine: Glyoxylate Aminotransferase 1 (AGT1) in M. oryzae and demonstrated that the AGT1 was indispensable for pathogenicity. The AGT1 knockout mutants were unable to penetrate the host plants, such as rice and barley, and therefore were non-pathogenic. The inability of ∆Moagt1 mutants to penetrate the susceptible plants was likely due to the disruption in coordination of the β-oxidation and the Glyoxylate cycle resulted from a blockage in lipid droplet mobilization and eventually utilization during conidial germination and appressorium morphogenesis, respectively. Here, we further demonstrate the role of AGT1 in lipid mobilization by in vitro germination assays and confocal microscopy.

  • peroxisomal Alanine Glyoxylate Aminotransferase agt1 is indispensable for appressorium function of the rice blast pathogen magnaporthe oryzae
    PLOS ONE, 2012
    Co-Authors: Vijai Bhadauria, Sabine Banniza, Albert Vandenberg, Gopalan Selvaraj, Yangdou Wei
    Abstract:

    The role of β-oxidation and the Glyoxylate cycle in fungal pathogenesis is well documented. However, an ambiguity still remains over their interaction in peroxisomes to facilitate fungal pathogenicity and virulence. In this report, we characterize a gene encoding an Alanine, Glyoxylate Aminotransferase 1 (AGT1) in Magnaporthe oryzae, the causative agent of rice blast disease, and demonstrate that AGT1 is required for pathogenicity of M. oryzae. Targeted deletion of AGT1 resulted in the failure of penetration via appressoria; therefore, mutants lacking the gene were unable to induce blast symptoms on the hosts rice and barley. This penetration failure may be associated with a disruption in lipid mobilization during conidial germination as turgor generation in the appressorium requires mobilization of lipid reserves from the conidium. Analysis of enhanced green fluorescent protein expression using the transcriptional and translational fusion with the AGT1 promoter and open reading frame, respectively, revealed that AGT1 expressed constitutively in all in vitro grown cell types and during in planta colonization, and localized in peroxisomes. Peroxisomal localization was further confirmed by colocalization with red fluorescent protein fused with the peroxisomal targeting signal 1. Surprisingly, conidia produced by the Δagt1 mutant were unable to form appressoria on artificial inductive surfaces, even after prolonged incubation. When supplemented with nicotinamide adenine dinucleotide (NAD+)+pyruvate, appressorium formation was restored on an artificial inductive surface. Taken together, our data indicate that AGT1-dependent pyruvate formation by transferring an amino group of Alanine to Glyoxylate, an intermediate of the Glyoxylate cycle is required for lipid mobilization and utilization. This pyruvate can be converted to non-fermentable carbon sources, which may require reoxidation of NADH generated by the β-oxidation of fatty acids to NAD+ in peroxisomes. Therefore, it may provide a means to maintain redox homeostasis in appressoria.

Vijai Bhadauria - One of the best experts on this subject based on the ideXlab platform.

  • Alanine: Glyoxylate Aminotransferase 1 is required for mobilization and utilization of triglycerides during infection process of the rice blast pathogen, Magnaporthe oryzae.
    Plant signaling & behavior, 2012
    Co-Authors: Vijai Bhadauria, Sabine Banniza, Albert Vandenberg, Gopalan Selvaraj, Yangdou Wei
    Abstract:

    The rice blast pathogen, Magnaporthe oryzae has been widely used as a model pathogen to study plant infection-related fungal morphogenesis, such as penetration via appressorium and plant-microbe interactions at the molecular level. Previously, we identified a gene encoding peroxisomal Alanine: Glyoxylate Aminotransferase 1 (AGT1) in M. oryzae and demonstrated that the AGT1 was indispensable for pathogenicity. The AGT1 knockout mutants were unable to penetrate the host plants, such as rice and barley, and therefore were non-pathogenic. The inability of ∆Moagt1 mutants to penetrate the susceptible plants was likely due to the disruption in coordination of the β-oxidation and the Glyoxylate cycle resulted from a blockage in lipid droplet mobilization and eventually utilization during conidial germination and appressorium morphogenesis, respectively. Here, we further demonstrate the role of AGT1 in lipid mobilization by in vitro germination assays and confocal microscopy.

  • peroxisomal Alanine Glyoxylate Aminotransferase agt1 is indispensable for appressorium function of the rice blast pathogen magnaporthe oryzae
    PLOS ONE, 2012
    Co-Authors: Vijai Bhadauria, Sabine Banniza, Albert Vandenberg, Gopalan Selvaraj, Yangdou Wei
    Abstract:

    The role of β-oxidation and the Glyoxylate cycle in fungal pathogenesis is well documented. However, an ambiguity still remains over their interaction in peroxisomes to facilitate fungal pathogenicity and virulence. In this report, we characterize a gene encoding an Alanine, Glyoxylate Aminotransferase 1 (AGT1) in Magnaporthe oryzae, the causative agent of rice blast disease, and demonstrate that AGT1 is required for pathogenicity of M. oryzae. Targeted deletion of AGT1 resulted in the failure of penetration via appressoria; therefore, mutants lacking the gene were unable to induce blast symptoms on the hosts rice and barley. This penetration failure may be associated with a disruption in lipid mobilization during conidial germination as turgor generation in the appressorium requires mobilization of lipid reserves from the conidium. Analysis of enhanced green fluorescent protein expression using the transcriptional and translational fusion with the AGT1 promoter and open reading frame, respectively, revealed that AGT1 expressed constitutively in all in vitro grown cell types and during in planta colonization, and localized in peroxisomes. Peroxisomal localization was further confirmed by colocalization with red fluorescent protein fused with the peroxisomal targeting signal 1. Surprisingly, conidia produced by the Δagt1 mutant were unable to form appressoria on artificial inductive surfaces, even after prolonged incubation. When supplemented with nicotinamide adenine dinucleotide (NAD+)+pyruvate, appressorium formation was restored on an artificial inductive surface. Taken together, our data indicate that AGT1-dependent pyruvate formation by transferring an amino group of Alanine to Glyoxylate, an intermediate of the Glyoxylate cycle is required for lipid mobilization and utilization. This pyruvate can be converted to non-fermentable carbon sources, which may require reoxidation of NADH generated by the β-oxidation of fatty acids to NAD+ in peroxisomes. Therefore, it may provide a means to maintain redox homeostasis in appressoria.

Yinai Zhang - One of the best experts on this subject based on the ideXlab platform.

  • metabolism of oxalate in humans a potential role kynurenine Aminotransferase glutamine transaminase cysteine conjugate beta lyase plays in hyperoxaluria
    Current Medicinal Chemistry, 2019
    Co-Authors: Cihan Yang, Jun Lu, Yinai Zhang, Jianyong Li
    Abstract:

    : Hyperoxaluria, excessive urinary oxalate excretion, is a significant health problem worldwide. Disrupted oxalate metabolism has been implicated in hyperoxaluria and accordingly, an enzymatic disturbance in oxalate biosynthesis can result in the primary hyperoxaluria. Alanine Glyoxylate Aminotransferase-1 and Glyoxylate reductase, the enzymes involving Glyoxylate (precursor for oxalate) metabolism, have been related to primary hyperoxalurias. Some studies suggest that other enzymes such as glycolate oxidase and Alanine Glyoxylate Aminotransferase-2 might be associated with primary hyperoxaluria as well, but evidence of a definitive link is not strong between the clinical cases and gene mutations. There are still some idiopathic hyperoxalurias, which require a further study for the etiologies. Some Aminotransferases, particularly kynurenine Aminotransferases, can convert Glyoxylate to glycine. Based on biochemical and structural characteristics, expression level, subcellular localization of some Aminotransferases, a number of them appear able to catalyze the transamination of Glyoxylate to glycine more efficiently than Alanine Glyoxylate Aminotransferase-1. The aim of this minireview is to explore other undermining causes of primary hyperoxaluria and stimulate research toward achieving a comprehensive understanding of underlying mechanisms leading to the disease. Herein, we reviewed all Aminotransferases in the liver for their functions in Glyoxylate metabolism. Particularly, kynurenine Aminotransferase-I and III were carefully discussed regarding their biochemical and structural characteristics, cellular localization, and enzyme inhibition. Kynurenine Aminotransferase-III is, so far, the most efficient putative mitochondrial enzyme to transaminate Glyoxylate to glycine in mammalian livers, might be an interesting enzyme to look over in hyperoxaluria etiology of primary hyperoxaluria and should be carefully investigated for its involvement in oxalate metabolism.

  • Metabolism of Oxalate in Humans: A Potential Role Kynurenine Aminotransferase/Glutamine Transaminase/Cysteine Conjugate Beta-lyase Plays in Hyperoxaluria.
    Current medicinal chemistry, 2019
    Co-Authors: Qian Han, Cihan Yang, Yinai Zhang
    Abstract:

    Hyperoxaluria, excessive urinary oxalate excretion, is a significant health problem worldwide. Disrupted oxalate metabolism has been implicated in hyperoxaluria and accordingly, an enzymatic disturbance in oxalate biosynthesis can result in the primary hyperoxaluria. Alanine Glyoxylate Aminotransferase-1 and Glyoxylate reductase, the enzymes involving Glyoxylate (precursor for oxalate) metabolism, have been related to primary hyperoxalurias. Some studies suggest that other enzymes such as glycolate oxidase and Alanine Glyoxylate Aminotransferase-2 might be associated with primary hyperoxaluria as well, but evidence of a definitive link is not strong between the clinical cases and gene mutations. There are still some idiopathic hyperoxalurias, which require a further study for the etiologies. Some Aminotransferases, particularly kynurenine Aminotransferases, can convert Glyoxylate to glycine. Based on biochemical and structural characteristics, expression level, subcellular localization of some Aminotransferases, a number of them appear able to catalyze the transamination of Glyoxylate to glycine more efficiently than Alanine Glyoxylate Aminotransferase-1. The aim of this minireview is to explore other undermining causes of primary hyperoxaluria and stimulate research toward achieving a comprehensive understanding of underlying mechanisms leading to the disease. Herein, we reviewed all Aminotransferases in the liver for their functions in Glyoxylate metabolism. Particularly, kynurenine Aminotransferase-I and III were carefully discussed regarding their biochemical and structural characteristics, cellular localization, and enzyme inhibition. Kynurenine Aminotransferase-III is, so far, the most efficient putative mitochondrial enzyme to transaminate Glyoxylate to glycine in mammalian livers, might be an interesting enzyme to look over in hyperoxaluria etiology of primary hyperoxaluria and should be carefully investigated for its involvement in oxalate metabolism.

Roman N. Rodionov - One of the best experts on this subject based on the ideXlab platform.

  • Overexpression of Alanine-Glyoxylate Aminotransferase 2 Protects from Asymmetric Dimethylarginine-induced Endothelial Dysfunction and Aortic Remodeling
    2021
    Co-Authors: Roman N. Rodionov, Dmitri Burdin, Vladimir T. Todorov, Natalia Jarzebska, Jens Martens-lobenhoffer, Anja Hofmann, Anne Kolouschek, Nada Cordasic, Johannes Jacobi, Elena Rubets
    Abstract:

    Abstract Objective: Elevated plasma concentrations of asymmetric dimethylarginine (ADMA) are associated with an increased risk of mortality and adverse cardiovascular outcomes. ADMA can be metabolized by dimethylarginine dimethylaminohydrolases (DDAHs) and by Alanine-Glyoxylate Aminotransferase 2 (AGXT2). Deletion of DDAH1 in mice leads to elevation of ADMA in plasma and blood pressure, while overexpression of human DDAH1 is associated with a lower plasma ADMA concentration and protective cardiovascular effects. The possible role of alternative metabolism of ADMA by AGXT2 remains to be elucidated. The goal of the current study was to test the hypothesis that transgenic overexpression of AGXT2 leads to lowering of plasma levels of ADMA and protection from vascular damage in the setting of DDAH1 deficiency.Approach and Results: We generated transgenic mice (TG) with ubiquitous overexpression of AGXT2. qPCR and Western Blot confirmed the expression of the transgene. Systemic ADMA levels were decreased by 15% in TG mice. In comparison with wild type animals plasma levels of asymmetric dimethylguanidino valeric acid (ADGV), the AGXT2 associated metabolite of ADMA, were six times higher. We crossed AGXT2 TG mice with DDAH1 knockout mice and observed that upregulation of AGXT2 lowers plasma ADMA and pulse pressure and protects the mice from endothelial dysfunction and adverse aortic remodeling.Conclusions: Upregulation of AGXT2 led to lowering of ADMA levels and protection from ADMA-induced vascular damage in the setting of DDAH1 deficiency. This is especially important, because all the efforts to develop pharmacological ADMA-lowering interventions by means of upregulation of DDAHs have been unsuccessful.

  • Abstract 453: Transgenic Overexpression of Alanine-Glyoxylate Aminotransferase 2 in Mice Lowers Asymmetric Dimethylarginine and Improves Vasomotor Function
    Arteriosclerosis Thrombosis and Vascular Biology, 2016
    Co-Authors: Natalia Jarzebska, Roman N. Rodionov, Dmitri Burdin, Silke Brilloff, Vladimir T. Todorov, Jens Martens-lobenhoffer, Anja Hofmann, Henning Morawietz, Anton V Demyanov, Karl F. Hilgers
    Abstract:

    Background: Elevation of the endogenous inhibitor of nitric oxide synthase asymmetric dimethylarginine (ADMA) has been shown to be associated with increased risk of cardiovascular diseases. There are two major pathways of ADMA catabolism: hydrolysis to citrulline by dimethylarginine dimethylaminohydrolases (DDAH) and transamination by Alanine-Glyoxylate Aminotransferase 2 (AGXT2) with formation of asymmetric dimethylguanidino valeric acid (ADGV). The second pathway is poorly characterized. The goal of the current study was to test the hypothesis that transgenic overexpression of AGXT2 leads to lowering of systemic levels of ADMA and improvement of vasomotor function. Methods and Results: We generated transgenic mice (TG) with ubiquitous overexpression of AGXT2 under control of the chicken beta actin (CAG) promoter. qPCR and Western Blot were used to confirm the ubiquitous expression of the transgene. There were no developmental or phenotypic changes in the TG animals. Biochemical data were generated using HPLC-MS/MS. ADMA plasma levels were decreased by 15% (p Conclusion: In the current study we demonstrated that upregulation of AGXT2 leads to lowering of ADMA levels and improvement of endothelium-dependent relaxation in vivo. AGXT2 thereby may be a potential drug target for long-term reduction of systemic ADMA levels in cardiovascular pathologies. This is especially important, because all the efforts to develop pharmacological ADMA-lowering interventions by means of upregulation of DDAH have not been successful so far.

  • Abstract 17603: Transgenic Overexpression of Alanine-Glyoxylate Aminotransferase 2 in Mice Lowers Asymmetric Dimethylarginine and Improves Vasomotor Function
    Circulation, 2015
    Co-Authors: Roman N. Rodionov, Dmitri Burdin, Silke Brilloff, Vladimir T. Todorov, Natalia Jarzebska, Jens Martens-lobenhoffer, Anja Hofmann, Henning Morawietz, Anton V Demyanov, Karl F. Hilgers
    Abstract:

    Background: ADMA (asymmetric dimethylarginine), an inhibitor of nitric oxide synthase, has been shown to be associated with the risk of cardiovascular diseases. There are two known pathways for ADMA catabolism: hydrolysis to citrulline by dimethylarginine dimethylaminohydrolases (DDAH) and transamination by Alanine-Glyoxylate Aminotransferase 2 (AGXT2) with formation of asymmetric dimethylguanidino valeric acid (ADGV). In contrast to the first pathway, the second one is poorly characterized. The goal of our study was to test the hypothesis that transgenic overexpression of AGXT2 would lead to lowering of systemic levels of ADMA and improvement of vasomotor function. Results: We generated transgenic mice (TG) with ubiquitous overexpression of AGXT2 under control of the chicken beta actin (CAG) promoter. qPCR and Western Blot were used to confirm the ubiquitous expression of the transgene. We did not observe and developmental or phenotypic abnormalities in the TG animals. Biochemical data were generated usi...

  • Abstract 373: Transgenic Overexpression of Alanine-Glyoxylate Aminotransferase 2 Lowers Tissue Levels of Asymmetric Dimethylarginine and Improves Endothelial Function in Mouse Aortas
    Arteriosclerosis Thrombosis and Vascular Biology, 2015
    Co-Authors: Roman N. Rodionov, Dmitri Burdin, Silke Brilloff, Vladimir T. Todorov, Natalia Jarzebska, Jens Martens-lobenhoffer, Anja Hofmann, Henning Morawietz, Anton V Demyanov, Renke Maas
    Abstract:

    Introduction: Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide synthase, which has been proposed to play a direct role in the pathogenesis of cardiovascular disease. There are two enzymatic pathways for degradation of ADMA: hydrolysis to citrulline by dimethylarginine dimethylaminohydrolase (DDAH) and transamination by Alanine-Glyoxylate Aminotransferase 2 (AGXT2) with formation of asymmetric dimethylguanidino valeric acid (ADGV). The first pathway is well characterized, whereas the physiological role of AGXT2 is still poorly understood. The goal of our study was to test the hypothesis that transgenic overexpression of AGXT2 would lead to lowering of systemic levels of ADMA and improved vasomotor function. Methods and results: We generated transgenic mice (TG) with ubiquitous overexpression of AGXT2 under control of the chicken beta actin (CAG) promoter. Ubiquitous overexpression of the transgene was confirmed by qPCR and Western Blot. TG animals had normal weight and no obser...

  • human Alanine Glyoxylate Aminotransferase 2 lowers asymmetric dimethylarginine and protects from inhibition of nitric oxide production
    Journal of Biological Chemistry, 2010
    Co-Authors: Roman N. Rodionov, Daryl J Murry, Sarah F Vaulman, Jeff W Stevens, Steven R Lentz
    Abstract:

    Elevated blood concentrations of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric-oxide (NO) synthase, are found in association with diabetes, hypertension, congestive heart failure, and atherosclerosis. ADMA levels are controlled by dimethylarginine dimethylaminohydrolases (DDAHs), cytosolic enzymes that hydrolyze ADMA to citrulline and dimethylamine. ADMA also has been proposed to be regulated through an alternative pathway by Alanine-Glyoxylate Aminotransferase 2 (AGXT2), a mitochondrial Aminotransferase expressed primarily in the kidney. The goal of this study was to define the subcellular localization of human AGXT2 and test the hypothesis that overexpression of human AGXT2 protects from ADMA-induced inhibition in nitric oxide (NO) production. AGXT2 was cloned from human kidney cDNA and overexpressed in COS-7 cells and human umbilical vein endothelial cells with a C-terminal FLAG epitope tag. Mitochondrial localization of human AGXT2 was demonstrated by confocal microscopy and a 41-amino acid N-terminal mitochondrial cleavage sequence was delineated by N-terminal sequencing of the mature protein. Overexpression of human AGXT2 in the liver of C57BL/6 mice using an adenoviral expression vector produced significant decreases in ADMA levels in plasma and liver. Overexpression of human AGXT2 also protected endothelial cells from ADMA-mediated inhibition of NO production. We conclude that mitochondrially localized human AGXT2 is able to effectively metabolize ADMA in vivo resulting in decreased ADMA levels and improved endothelial NO production.