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Leonor C. San Martín De Viale - One of the best experts on this subject based on the ideXlab platform.

  • Porphyrinogen carboxy-lyase from chick embryo liver—in vivo effect of heptachlor and lindane
    International Journal of Biochemistry, 2003
    Co-Authors: María Cristina Taira, Leonor C. San Martín De Viale
    Abstract:

    Abstract 1. 1. Heptachlor and lindane are widespread chlorinated insecticides. But porphyrinogen carboxy-lyase seems to be a key enzyme in the porphyria produced by chlorinated compounds in mammals. It was then considered interesting to perform the present investigation, the purpose of which was to study: (a) the porphyrinogenic ability of heptachlor and lindane when they were administered in vivo to whole chick embryo and (b) the in vivo effect of heptachlor and lindane on porphyrinogen carboxy-lyase activity. 2. 2. The results obtained showed that both compounds produce a significant accumulation of hepatic porphyrins, that produced by lindane being greater than that produced by heptachlor. 3. 3. The effect of heptachlor and lindane administration on porphyrinogen carboxy-lyase was studied in embryos from New Hampshire and Leghorn strains and in New Hampshire 1-day old chicks. The influence of phenobarbital and SKF-525 A pretreatment on the response of porphyrinogen carboxy-lyase to these drugs was also studied in New Hampshire chick embryos. The results indicate that the drugs do not produce decreased enzyme activity in the New Hampshire embryos, while a decrease of enzyme activity in the White Leghorn embryos and in the New Hampshire new-born chicks was observed. Results from pretreatment assays suggest that the decrease of porphyrinogen carboxy-lyase activity seems to be associated with the formation of a drug metabolite. 4. 4. It is concluded that heptachlor and lindane are porphyrinogenic drugs and that the response of decarboxylating enzyme to them seems to be dependent on: (a) the strain, (b) the development of the animal and (c) the drug metabolization system; some of the lindane and heptachlor metabolites being responsible for hepatic porphyrinogen carboxy-lyase decrease.

  • Porphyria-induced hepatic porphyrinogen carboxy-lyase inhibitor and its interaction with the active site(s) of the enzyme.
    Iubmb Life, 1999
    Co-Authors: Silvia Cristina Billi De Catabbi, R. Wainstok De Cahmanovici, Carolina Minutolo, Carmen Aldonatti, Leonor C. San Martín De Viale
    Abstract:

    Porphyrinogen carboxy-lyase is an enzyme that sequentially decarboxylates uroporphyrinogen III (8-COOH) to yield coproporphyrinogen III (4-COOH). In mammals this enzyme activity is impaired by hexachlorobenzene treatment, through generation of an enzyme inhibitor. The interaction of porphyrinogen carboxy-lyase inhibitor, extracted from the liver of hexachlorobenzene-treated rats, with substrate decarboxylation sites on the enzyme, was studied using four different carboxylated substrates belonging to the isomeric III series of naturally-formed porphyrinogens containing 8-,7-,6- and 5-COOH. Similar inhibitor effects were elicited against all the substrates assayed, with the exception of pentacarboxyporphyrinogen III in which decarboxylation was not inhibited to same extent. Enzyme protection assays in the presence of the different substrates, indicated that each porphyrinogen protects its own decarboxylation from inhibitor action. Preincubation of the inhibitor with normal enzyme increased its inhibitory effect. On the other hand, preincubation of both enzyme and inhibitor with superoxide dismutase or mannitol, did not alter inhibitory activity. Preincubation of the inhibitor with a number of aminoacids showed that only arginine and its derivative Nα-Benzoyl-L-Arginine ethyl ester interact with the inhibitor, noticeably reducing its ability to inhibit porphyrinogen carboxy-lyase. Albumin, histidine, serine, cysteine and imidazol, were unable to quench inhibitor activity. The present results indicate that the inhibitor acts at the binding site of each porphyrinogen. Taking into account that arginine is related to enzyme activity, and that histidine is found at the binding site of the substrates, the results suggest that the inhibitor could bind to arginine residues, blocking the access of substrates to histidine and altering the adequate orientation for decarboxylation by masking the positively charged active site necessary for porphyrinogen binding to the enzyme. In addition an indirect effect of the inhibitor mediated through free radicals could be discarded.

Toru Matoh - One of the best experts on this subject based on the ideXlab platform.

  • Purification and cDNA cloning of UDP-D-glucuronate carboxy-lyase (UDP-D-xylose synthase) from pea seedlings.
    Plant & cell physiology, 2002
    Co-Authors: Masaru Kobayashi, Hironobu Nakagawa, Izumi Suda, Isao Miyagawa, Toru Matoh
    Abstract:

    Uridine diphospho-D-glucuronate carboxy-lyase (UDP-D-xylose synthase; EC 4.1.1.35), which catalyzes the conversion of UDP-D-glucuronate to UDP-D-xylose, was purified to apparent homogenity from pea (Pisum sativum L.) seedlings. The pH optimum for enzyme activity was around 5-6, and the activity was not affected by exogeneously supplied NAD + and NADH. The purified enzyme had a molecular weight of 250 kDa and consisted of 42 kDa polypeptides. Based on the amino acid sequence, a probe (400 bp) was prepared with degenerate primers by a reverse transcriptase-PCR. Using this probe, a clone encoding 346 amino acid residues was screened from a pea cDNA library. The recombinant protein expressed in Escherichia coli catalyzed conversion of UDP-D-glucuronate to UDP-D-xylose, confirming that the isolated clone encoded UDP-D-glucuronate carboxy-lyase.

Masaru Kobayashi - One of the best experts on this subject based on the ideXlab platform.

  • Purification and cDNA cloning of UDP-D-glucuronate carboxy-lyase (UDP-D-xylose synthase) from pea seedlings.
    Plant & cell physiology, 2002
    Co-Authors: Masaru Kobayashi, Hironobu Nakagawa, Izumi Suda, Isao Miyagawa, Toru Matoh
    Abstract:

    Uridine diphospho-D-glucuronate carboxy-lyase (UDP-D-xylose synthase; EC 4.1.1.35), which catalyzes the conversion of UDP-D-glucuronate to UDP-D-xylose, was purified to apparent homogenity from pea (Pisum sativum L.) seedlings. The pH optimum for enzyme activity was around 5-6, and the activity was not affected by exogeneously supplied NAD + and NADH. The purified enzyme had a molecular weight of 250 kDa and consisted of 42 kDa polypeptides. Based on the amino acid sequence, a probe (400 bp) was prepared with degenerate primers by a reverse transcriptase-PCR. Using this probe, a clone encoding 346 amino acid residues was screened from a pea cDNA library. The recombinant protein expressed in Escherichia coli catalyzed conversion of UDP-D-glucuronate to UDP-D-xylose, confirming that the isolated clone encoded UDP-D-glucuronate carboxy-lyase.

Silvia Cristina Billi De Catabbi - One of the best experts on this subject based on the ideXlab platform.

  • Porphyria-induced hepatic porphyrinogen carboxy-lyase inhibitor and its interaction with the active site(s) of the enzyme.
    Iubmb Life, 1999
    Co-Authors: Silvia Cristina Billi De Catabbi, R. Wainstok De Cahmanovici, Carolina Minutolo, Carmen Aldonatti, Leonor C. San Martín De Viale
    Abstract:

    Porphyrinogen carboxy-lyase is an enzyme that sequentially decarboxylates uroporphyrinogen III (8-COOH) to yield coproporphyrinogen III (4-COOH). In mammals this enzyme activity is impaired by hexachlorobenzene treatment, through generation of an enzyme inhibitor. The interaction of porphyrinogen carboxy-lyase inhibitor, extracted from the liver of hexachlorobenzene-treated rats, with substrate decarboxylation sites on the enzyme, was studied using four different carboxylated substrates belonging to the isomeric III series of naturally-formed porphyrinogens containing 8-,7-,6- and 5-COOH. Similar inhibitor effects were elicited against all the substrates assayed, with the exception of pentacarboxyporphyrinogen III in which decarboxylation was not inhibited to same extent. Enzyme protection assays in the presence of the different substrates, indicated that each porphyrinogen protects its own decarboxylation from inhibitor action. Preincubation of the inhibitor with normal enzyme increased its inhibitory effect. On the other hand, preincubation of both enzyme and inhibitor with superoxide dismutase or mannitol, did not alter inhibitory activity. Preincubation of the inhibitor with a number of aminoacids showed that only arginine and its derivative Nα-Benzoyl-L-Arginine ethyl ester interact with the inhibitor, noticeably reducing its ability to inhibit porphyrinogen carboxy-lyase. Albumin, histidine, serine, cysteine and imidazol, were unable to quench inhibitor activity. The present results indicate that the inhibitor acts at the binding site of each porphyrinogen. Taking into account that arginine is related to enzyme activity, and that histidine is found at the binding site of the substrates, the results suggest that the inhibitor could bind to arginine residues, blocking the access of substrates to histidine and altering the adequate orientation for decarboxylation by masking the positively charged active site necessary for porphyrinogen binding to the enzyme. In addition an indirect effect of the inhibitor mediated through free radicals could be discarded.

Marie E Fraser - One of the best experts on this subject based on the ideXlab platform.

  • identification of the active site residues in atp citrate lyase s carboxy terminal portion
    Protein Science, 2019
    Co-Authors: Vinh Nguyen, Noreen Singh, Ana Medina, Isabel Uson, Marie E Fraser
    Abstract:

    ATP-citrate lyase (ACLY) catalyzes production of acetyl-CoA and oxaloacetate from CoA and citrate using ATP. In humans, this cytoplasmic enzyme connects energy metabolism from carbohydrates to the production of lipids. In certain bacteria, ACLY is used to fix carbon in the reductive tricarboxylic acid cycle. The carboxy(C)-terminal portion of ACLY shows sequence similarity to citrate synthase of the tricarboxylic acid cycle. To investigate the roles of residues of ACLY equivalent to active site residues of citrate synthase, these residues in ACLY from Chlorobium limicola were mutated, and the proteins were investigated using kinetics assays and biophysical techniques. To obtain the crystal structure of the C-terminal portion of ACLY, full-length C. limicola ACLY was cleaved, first non-specifically with chymotrypsin and subsequently with Tobacco Etch Virus protease. Crystals of the C-terminal portion diffracted to high resolution, providing structures that show the positions of active site residues and how ACLY tetramerizes.