delta-Aminolevulinic acid synthase from Euglena gracilis.

Beale, Samuel I.; Foley, Terrence; Dzelzkalns, Valdis A.

doi:10.1073/pnas.78.3.1666

Cited by 52 publications

(37 citation statements)

References 24 publications

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“…These questions concern yet another route, that of the 'classical' ALA synthase which has recently been demonstrated in Euglena (2), and indications for its operation were obtained also from etiolated leaves (26). The latter enzyme may take part in porphyrin synthesis in the dark or outside the chloroplast (2,26).…”

Section: Discussionmentioning

confidence: 99%

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Alternative Routes for the Synthesis of 5-Aminolevulinic Acid in Maize Leaves

Harel¹,

Ne’eman²,

Meller³

1983

Plant Physiol.

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Cell-free extracts from greening maize (Zea mays L.) leaves catalyze the conversion of I'4C]2-ketoglutarate (KG) to 14C15-aminolevulinic acid (ALA) in a reaction which requires NADH and an amino donor and shows maximal activity around pH 6.5. The enzymic system is located in the cytosol. This cell fraction contains a low level of 'KG dehydrogenase' activity and a transaminase which catalyzes the conversion of 4,5-dioxovaleric acid (DOVA) to ALA. The transaminase can use glutamate, aspartate, or alanine as amino donor. It is effectively inhibited by aminooxyacetate and ethylenediamine tetraacetate and shows maximal activity at pH 6.7. The activity of DOVA transaminase is only sllghdy affected by preillumination of leaves and can also be detected in green leaves and in roots.DOVA was isolated from leaves and roots and determined as its benzoquinoxaline derivative. Significant amounts were found only in tissues in which ALA had accumulated or after it was exogenously supplied.DOVA was labeled in vivo by both 114CIALA and I14CIKG. Small amounts were also formed from ALA in a cell-free system. It is suggested that DOVA may be an intermediate in the diversion of ALA to respiratory metabollsm and that it is not involved in the biosynthesis of this porphyrin precursor.

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Section: Discussionmentioning

confidence: 99%

“…The 'DOVA route' might be involved in ALA formation in the dark, responsible for the low level of Pchlide found in etiolated leaves (11) or for the synthesis of extra plastidial porphyrins. However, since the latter function is apparently carried out by the classical ALA synthase (2,26), the DOVA route might perform yet another function.…”

Section: Discussionmentioning

confidence: 99%

Alternative Routes for the Synthesis of 5-Aminolevulinic Acid in Maize Leaves

Harel¹,

Ne’eman²,

Meller³

1983

Plant Physiol.

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“…It was recently reported that Euglena gracilis contains ALA synthase (6). High levels of ALA synthase activity were measured in extracts of aplastidic mutant cells and dark-grown wild-type cells, and lower levels were found in light-grown wild-type cells.…”

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confidence: 99%

“…Cells were grown at 23°C and 32 ltE m-2s-' of cool white and red fluorescent light in a glucose-based heterotrophic medium as previously described (6). Cell population densities were determined with a Coulter Counter (Model ZBI, Coulter Electronics).…”

mentioning

confidence: 99%

“…Chl was extracted into methanol and quantitated by spectrophotometry, using the absorption coefficients of MacKinney (19). ALA synthase was extracted and assayed by the methods previously reported (6), except that cells were disrupted by sonication for four periods using a Sonifier (Model W 185, Heat Systems-Ultrasonics), and ALA was measured by condensation with ethyl acetoacetate at pH 6.8 (18) followed by color development with Ehrlich Hg reagent (26). Spectrophotometry was performed on a Cary Model 219 instrument (Varian).…”

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Induction of δ-Aminolevulinic Acid Synthase Activity and Inhibition of Heme Synthesis in Euglena gracilis by N-Methyl Mesoporphyrin IX

Beale

Foley²

1982

Plant Physiol.

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N-Methyl mesoporphyrin IX, an inhibitor of heme synthesis, increases extractable 8-aminolevulinic acid (ALA) synthase activity when administered to growing cultures of Euglena gracilis Klebs strain Z Pringsheim in micromolar concentrations. Wild-type light-grown green cells and white aplastidic cells exhibited 2.8-fold and 1.8-fold increases, respectively, in ALA synthase activity within five to six hours after incubation with 4 x 106 molar N-methyl mesoporphyrin IX. Protoheme levels were decreased and 59Fe incorporation into heme was inhibited by N-methyl mesoporphyrin IX, indicating that, as in animal cells, N-methyl mesoporphyrin IX acts specifically to block iron insertion into protoporphyrin IX. Chlorophyll synthesis in wild-type cells was not affected within the first 6 hours after administration of N-methyl mesoporphyrin IX. In animals, fungi, and bacteria, the first identified step hepatic ALA synthase within 4 h after injection into mice. We now report that N-methyl mesoporphyrin IX increases extractable ALA synthase activity, inhibits incorporation of exogenous 59Feinto heme in vivo, and diminishes heme levels in Euglena, but has no effect on Chl formation within the first 6 h after administration.MATERIALS AND METHODS Cultures of Euglena gracilis Klebs strain Z Pringsheim and an aplastidic mutant derived from this strain, W14ZNa 1 L (23), were kindly provided by H. Lyman (State University of New York, Stony Brook, NY). Cells were grown at 23°C and 32 ltE m-2s-' of cool white and red fluorescent light in a glucose-based heterotrophic medium as previously described (6). Cell population densities were determined with a Coulter Counter (Model ZBI, Coulter Electronics). Chl was extracted into methanol and quantitated by spectrophotometry, using the absorption coefficients of MacKinney (19). ALA synthase was extracted and assayed by the methods previously reported (6), except that cells were disrupted by sonication for four periods using a Sonifier (Model W 185, Heat Systems-Ultrasonics), and ALA was measured by condensation with ethyl acetoacetate at pH 6.8 (18) followed by color development with Ehrlich Hg reagent (26). Spectrophotometry was performed on a Cary Model 219 instrument (Varian).Protoheme was extracted by slight modification of the method of Stillman and Gassman (25). All operations were carried out at ice temperature. Cells were extracted with three 5-ml portions of 90% acetone containing 10-2 M NH40H. These extracts were discarded. The cells were then extracted with 2 ml acetone containing 2% HCI. A second l-ml acetone-HCl extraction supernatant was added to the first, and then 2 ml peroxide-free diethyl ether were added to the combined acetone extracts. After mixing, 6 ml H20 were added, the solution was thoroughly mixed, and briefly centrifuged to separate the phases. The upper ether phase was removed and the lower phase was extracted with I ml ether. The combined heme-containing ether phases were backwashed once with 1 ml H20. After evaporation of the ether, protoheme was de...

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δ‐Aminolevulinic acid biosynthesis in Ustilago maydis

Schneegurt

2005

J. Basic Microbiol.

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A biosynthetic precursor of tetrapyrroles, δ-aminolevulinic acid (ALA), can be formed via two pathways: enzymatic condensation of glycine and succinyl-CoA by ALA synthase in animal mitochondria and some fungi, and the C 5 pathway converting glutamate to ALA in plants, algae, archaea, and most bacteria. The two pathways are distinguishable using specifically radiolabeled compounds. The C 1 of glutamate is lost during conversion to succinate in the TCA cycle, and the C 2 of glycine is lost during conversion to acetyl-CoA on the way to glutamate. Desalted high-speed supernatants of Ustilago maydis sporidia extracts were assayed using specifically radiolabeled substrates. A significant amount of radiolabel was incorporated into ALA from 2-[ 14 C]glycine. No radiolabel was incorporated into ALA from 1-[ 14 C]glutamate. These results indicate that the basidiomycete yeast, Ustilago maydis, has active ALA synthase.

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delta-Aminolevulinic acid synthase from Euglena gracilis.

Cited by 52 publications

References 24 publications

Alternative Routes for the Synthesis of 5-Aminolevulinic Acid in Maize Leaves

Alternative Routes for the Synthesis of 5-Aminolevulinic Acid in Maize Leaves

Induction of δ-Aminolevulinic Acid Synthase Activity and Inhibition of Heme Synthesis in Euglena gracilis by N-Methyl Mesoporphyrin IX

δ‐Aminolevulinic acid biosynthesis in Ustilago maydis

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