Isolation and characterization of the gene coding for the amidinotransferase involved in the biosynthesis of phaseolotoxin in Pseudomonas syringae pv. phaseolicola. (1/58)

Pseudomonas syringae pv. phaseolicola is the causal agent of the "halo blight" disease of beans. A key component in the development of the disease is a nonhost-specific toxin, Ndelta-(N'-sulphodiaminophosphinyl)-ornithyl-alanyl-homoarginine, known as phaseolotoxin. The homoarginine residue in this molecule has been suggested to be the product of L-arginine:lysine amidinotransferase activity, previously detected in extracts of P. syringae pv. phaseolicola grown under conditions of phaseolotoxin production. We report the isolation and characterization of an amidinotransferase gene (amtA) from P. syringae pv. phaseolicola coding for a polypeptide of 362 residues (41.36 kDa) and showing approximately 40% sequence similarity to L-arginine:inosamine-phosphate amidinotransferase from three species of Streptomyces spp. and 50.4% with an L-arginine:glycine amidinotransferase from human mitochondria. The cysteine, histidine, and aspartic acid residues involved in substrate binding are conserved. Furthermore, expression of the amtA and argK genes and phaseolotoxin production occurs at 18 degrees C but not at 28 degrees C. An amidinotransferase insertion mutant was obtained that lost the capacity to synthesize homoarginine and phaseolotoxin. These results show that the amtA gene isolated is responsible for the amidinotransferase activity detected previously and that phaseolotoxin production depends upon the activity of this gene.  (+info)

Arginine:glycine amidinotransferase deficiency: the third inborn error of creatine metabolism in humans. (2/58)

Arginine:glycine amidinotransferase (AGAT) catalyzes the first step of creatine synthesis, resulting in the formation of guanidinoacetate, which is a substrate for creatine formation. In two female siblings with mental retardation who had brain creatine deficiency that was reversible by means of oral creatine supplementation and had low urinary guanidinoacetate concentrations, AGAT deficiency was identified as a new genetic defect in creatine metabolism. A homozygous G-A transition at nucleotide position 9297, converting a tryptophan codon (TGG) to a stop codon (TAG) at residue 149 (T149X), resulted in undetectable cDNA, as investigated by reverse-transcription PCR, as well as in undetectable AGAT activity, as investigated radiochemically in cultivated skin fibroblasts and in virus-transformed lymphoblasts of the patients. The parents were heterozygous for the mutant allele, with intermediate residual AGAT activities. Recognition and treatment with oral creatine supplements may prevent neurological sequelae in affected patients.  (+info)

Methylation demand and homocysteine metabolism: effects of dietary provision of creatine and guanidinoacetate. (3/58)

S-adenosylmethionine, formed by the adenylation of methionine via S-adenosylmethionine synthase, is the methyl donor in virtually all known biological methylations. These methylation reactions produce a methylated substrate and S-adenosylhomocysteine, which is subsequently metabolized to homocysteine. The methylation of guanidinoacetate to form creatine consumes more methyl groups than all other methylation reactions combined. Therefore, we examined the effects of increased or decreased methyl demand by these physiological substrates on plasma homocysteine by feeding rats guanidinoacetate- or creatine-supplemented diets for 2 wk. Plasma homocysteine was significantly increased (~50%) in rats maintained on guanidinoacetate-supplemented diets, whereas rats maintained on creatine-supplemented diets exhibited a significantly lower (~25%) plasma homocysteine level. Plasma creatine and muscle total creatine were significantly increased in rats fed the creatine-supplemented or guanidinoacetate-supplemented diets. The activity of kidney L-arginine:glycine amidinotransferase, the enzyme catalyzing the synthesis of guanidinoacetate, was significantly decreased in both supplementation groups. To examine the role of the liver in mediating these changes in plasma homocysteine, isolated rat hepatocytes were incubated with methionine in the presence and absence of guanidinoacetate and creatine, and homocysteine export was measured. Homocysteine export was significantly increased in the presence of guanidinoacetate. Creatine, however, was without effect. These results suggest that homocysteine metabolism is sensitive to methylation demand imposed by physiological substrates.  (+info)

On the control of arginine metabolism in chicken kidney and liver. (4/58)

Arginases have been found to be located on the external side of the inner mitochondrial membrane of chicken kidney and liver. Transamidinase has been detected within the liver mitochondrial matrix space. Arginases and transamidinase act upon two different intracellular arginine pools. Penetration of arginine into the matrix space occurs only in respring mitochondria and in the presence of anions such as acetate and phosphate; D-arginine, L-ornithine, D-'ornithine and L-lysine penetrate with the same modalities. L-Histidine penetrates only kidney mitochondria. Because of transamidinase compartmentation, the rate of creatine synthesis is influenced by the rate of penetration of arginine into the mitochondria.  (+info)

Guanidinoacetate and creatine plus creatinine assessment in physiologic fluids: an effective diagnostic tool for the biochemical diagnosis of arginine:glycine amidinotransferase and guanidinoacetate methyltransferase deficiencies. (5/58)

BACKGROUND: Disorders of creatine metabolism arise from genetic alterations of arginine:glycine amidinotransferase (AGAT), guanidinoacetate methyltransferase (GAMT), and the creatine transporter. We developed a strategy for the detection of AGAT and GAMT defects by measurement of guanidinoacetate (GAA) and creatine plus creatinine (Cr+Crn) in biological fluids. METHODS: Three patients with AGAT deficiency from the same pedigree and their eight relatives, as well as a patient affected by a GAMT defect and his parents were analyzed by a new HPLC procedure in comparison with 90 controls. The method, which uses precolumn derivatization with benzoin, separation with a reversed-phase column, and fluorescence detection, has shown good precision and sensitivity and requires minimal sample handling. RESULTS: In the three AGAT patients, plasma GAA was 0.01-0.04 micro mol/L [mean (SD) for neurologically normal controls was 1.16 (0.59) micromol/L], Cr+Crn was 15-29 micro mol/L [reference limit in our laboratory, 79 (38) micromol/L]. Urinary GAA was 2.4-5.8 micro mol/L [reference, 311 (191) micromol/L], and Cr+Crn was 2.1-3.3 mmol/L [reference, 9.9 (4.1) mmol/L]. We found a smaller decrease in GAA and Cr+Crn in some carriers of an AGAT defect. In the patient with GAMT deficiency, plasma and urine GAA was increased (18.6 and 1783 micromol/L, respectively), and Cr+Crn was decreased in plasma (10.7 micromol/L) and urine (2.1 mmol/L). GAA was increased in the parents' plasmas and in the mother's urine. CONCLUSION: The assessment of GAA is a new tool for the detection of both GAMT and AGAT deficiencies.  (+info)

Fresh and cultured thyroid gland: survival and function after implantation. (6/58)

Isogeneic or allogeneic thyroid glands were implanted into thyroidectomized recipeint rats. These grafts, either fresh or cultured, were placed in hamstring muscle pocket or under the renal capsule. Survival and function of the grafts were evaluated by: restoration of normal levels of serum thyroxine, weight gain, kidney transamidinase (a thyroxine-induced enzyme), and histological appearance of re-excised implants. A isografts, fresh and cultured, functioned well as ectopic thyroid glands, though restoration of normal serum thyroxine levels was more rapid for the fresh implants. Fresh allografts functioned transiently but then failed due to rejection. No function was detected for cultured allografts, and rejection was seen histologically. The rat thyroid allograft therefore differs from the rat parathyroid allograft, which often can function for several months despite histological evidence of rejection. Maintenance in tissue culture prior to implantation does not appear to alter the longterm immunogenicity of either organ.  (+info)

Gatm, a creatine synthesis enzyme, is imprinted in mouse placenta. (7/58)

To increase our understanding of imprinting and epigenetic gene regulation, we undertook a search for new imprinted genes. We identified Gatm, a gene that encodes l-arginine:glycine amidinotransferase, which catalyzes the rate-limiting step in the synthesis of creatine. In mouse, Gatm is expressed during development and is imprinted in the placenta and yolk sac, but not in embryonic tissues. The Gatm gene maps to mouse chromosome 2 in a region not previously shown to contain imprinted genes. To determine whether Gatm is located in a cluster of imprinted genes, we investigated the expression pattern of genes located near Gatm: Duox1-2, Slc28a2, Slc30a4 and a transcript corresponding to LOC214616. We found no evidence that any of these genes is imprinted in placenta. We show that a CpG island associated with Gatm is unmethylated, as is a large CpG island associated with a neighboring gene. This genomic screen for novel imprinted genes has elucidated a new connection between imprinting and creatine metabolism during embryonic development in mammals.  (+info)

Studies on the catabolism of Ng-methylarginine, Ng, Ng-dimethylarginine and Ng, Ng-dimethylarginine in the rabbit. (8/58)

1. The routes of elimination of Ng-methylarginine, Ng, Ng-dimethylarginine and Ng, Ng-dimethylarginine were investigated in the rabbit. 2. Analyses showed low plasma concentrations of these amino acids (around 1 nmol/ml) and ratios similar to those found in tissue proteins. The concentrations of these amino acids in extracts of brain, kidney, liver and spleen were similar except that liver had a lower concentration of Ng-methylarginine and Ng, Ng-dimethylarginine. Cerebrospinal fluid contained traces of each amino acid.  (+info)

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