δ-Aminolevulinic acid synthase of Euglena gracilis: Physical and kinetic properties

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

doi:10.1016/0003-9861(82)90204-1

Cited by 21 publications

(7 citation statements)

References 30 publications

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“…A study of the physical and kinetic properties of the ALA synthase of Euglena gracilis revealed that it is similar to ALA synthase isolated from other sources with respect to molecular weight, pH optimum, and substrate affinities (8). The regulatory behavior of ALA synthase in Euglena reported here provides information concerning its role in this organism.…”

supporting

confidence: 56%

“…The broken cell suspensions were centrifuged in the microcentrifuge for 1 min at 13,000g to yield supernatant for the ALA synthase assay. This extraction procedure consistently yielded ALA synthase with greater activity per number of extracted cells than the previously used procedure (4,8).…”

mentioning

confidence: 73%

“…ALA synthase isolated from animal and bacteria sources has been reported to be inhibited by micromolar concentrations of hemes and porphyrins (6, 21, 29, 31). Because ALA synthase from Euglena has physical (8), the possibility of allosteric inhibition by hemin, protoporphyrin IX, and PBG was examined at concentrations up to 100/,M in extracts of aplastidic cells that had been partially purified by passage through a column of Sephadex G-25 in extraction medium to remove low molecular weight components. None of these potential allosteric feedback inhibitors had a pronounced effect on ALA synthase activity within the range of concentrations examined (Table V).…”

mentioning

confidence: 99%

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δ-Aminolevulinic Acid Synthase of Euglena gracilis: Regulation of Activity

Foley¹,

Dzelzkalns²,

Beale³

1982

Plant Physiol.

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ABSTRACTregions, which thereby supplants the necessity for ALA formation via the ALA synthase route.8-Aminolevulinic acid (ALA), a key precursor of the tetrapyrroles heme and chlorophyll, is capable of being synthesized by two different routes in cells of the unicellular green alga Euglena gracilis: from the intact carbon skeleton of glutamate, and via the condensation of glycine and succinyl CoA, mediated by the enzyme ALA synthase. The regulatory properties of ALA synthase were examined in order to establish its role in Euglena.Partially purified Euglena ALA synthase, unlike the case with the bacterial or animal-derived enzyme, does not exhibit allosteric inhibition by the tetrapyrrole pathway products heme, protoporphyrin IX, and porphobilinogen, at concentrations up to 100 micromolar.In aplastidic mutant cells, extractable ALA synthase activity is constant during exponential growth, and decreases to low levels as the cells reach the stationary state. Rapid exponential decline of ALA synthase (t1/2 = 55 min) occurs after administration of 43 micromolar cycloheximide, but not 6.2 millimolar chloramphenicol. These results suggest that, as in other eukaryotic cells, ALA synthase is synthesized on cytoplasmic ribosomes and is subject to rapid turnover in vivo.Extractable ALA synthase activity increases 2.5-fold within 6 hours after administration of 100 millimolar ethanol, a stimulator of mitochondrial development, and 4.5-fold within 12 hours after administration of I millimolar 4,6-dioxoheptanoic acid, which blocks ALA utilization, suggesting that activity is controlled in vivo by a feedback induction-repression mechanism, coupled with rapid enzyme turnover.In heterotrophically grown wild-type cells, low levels of ALA synthase rapidly increase 4.5-fold within 12 hours after cells are transferred from the light to the dark, and decrease exponentially (t1/2 = 75 min) when cells are transferred from the dark to light. The dark levels are equal to those in light-or dark-grown aplastidic mutant cells. The low level occurring in light-grown wild-type cells is not altered by the presence of 10 micromolar 3-(3,4-dichlorophenyl)-1,1-dimethylurea, which blocks photosynthetic 02 production. The decrease that occurs on dark-to-light transfer can be diminished by 12-or 24-hour prior incubation with 6.2 millimolar chloramphenicol, which also retards chlorophyll synthesis after the transfer to light.The positive relationship of ALA synthase activity to degree of mitochondrial expression, and the inverse relationship to plastid development and chlorophyll synthesis, suggests that ALA synthase functions to provide precursors to nonplastid tetrapyrroles in Euglena. In light-grown, wild-type cells, the diminished levels of ALA synthase may be due to the ability of developing plastids to export heme or a heme precursor to other cellular '

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confidence: 56%

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confidence: 73%

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confidence: 99%

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δ-Aminolevulinic Acid Synthase of Euglena gracilis: Regulation of Activity

Foley¹,

Dzelzkalns²,

Beale³

1982

Plant Physiol.

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“…[2][3][4]24,36 Examination of the physical and kinetic properties of the E. gracilis ALAS and the regulation of its activity indicate that ALAS exclusively functions in E. gracilis to provide precursors for non-plastid tetrapyrroles. 4,37 This conclusion was subsequently proven directly by measuring incorporation of labeled precursors into isolated tetrapyrroles specific to plastids and mitochondria. 5 Under all growth conditions tested, heme a of mitochondrial cytochrome c oxidase was formed from glycine, while in cells growing under conditions permitting chlorophyll formation, the plastid pigments were formed exclusively from glutamate.…”

Section: B C 4 (Shemin) and C 5 Pathwaysmentioning

confidence: 95%

Toward Heme: 5-Aminolevulinate Synthase and Initiation of Porphyrin Synthesis

Fratz¹,

Stojanovski²,

Ferreira³

2013

Handbook of Porphyrin Science

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“…Two major pathways for the biosynthesis of 5-aminolevulinic acid have been described. The biosynthesis of 5-aminolevulinic acid in bacterial cells is tightly regulated by feedback inhibition at the level of 5-aminolevulinic acid formation, and 5-aminolevulinic acid formation is considered to be the ratelimiting step for tetrapyrrole biosynthesis (Dzelzkalns et al 1992).…”

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confidence: 99%

Expression of 5-aminolevulinic acid synthase in recombinant Escherichia coli

Wang

Zhang

2006

World J Microbiol Biotechnol

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The Rhodobacter sphaeroides hemA gene codes for 5-aminolevulinic acid synthase (EC.2.3.1.3), catalysing the pyridoxal phosphate-dependent condensation of succinyl coenzyme A and glycine to give 5-aminolevulinic acid. The gene was transformed into Escherichia coli K 12 using the pALA vector systems. Effects of host strains, vector plasmids, growth substrates and precursors on the expression and activity of 5-aminolevulinic acid synthase were studied. The E. coli host strain had an enormous effect on 5-aminolevulinic acid synthase activity and production of 5-aminolevulinic acid, with E. coli DH1 being best suited. RT-PCR of hemA mRNA indicated that the transcription of the hemA gene in the recombinant strain appeared to be higher than that of the wild-type strain. 5-Aminolevulinic acid synthase activity was maximal when hemA had the same transcription direction as the lac promoter. Distance between lac promoter and hemA affected the expression of 5-aminolevulinic acid synthase on different growth substrates. 5-Aminolevulinic acid synthase activities were also dependent on the carbon sources and precursors. L-Malate gave the highest activity of 5-aminolevulinic acid synthase, while lactose as a carbon source resulted in a repression of 5-aminolevulinic acid synthase. Production of 5-aminolevulinic acid by recombinants were limited by the availability of glycine, and repeated addition of glycine (20 mM) to the growth medium increased production of 5-aminolevulinic acid to 33.8 mM.

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δ-Aminolevulinic acid synthase of Euglena gracilis: Physical and kinetic properties

Cited by 21 publications

References 30 publications

δ-Aminolevulinic Acid Synthase of Euglena gracilis: Regulation of Activity

δ-Aminolevulinic Acid Synthase of Euglena gracilis: Regulation of Activity

Toward Heme: 5-Aminolevulinate Synthase and Initiation of Porphyrin Synthesis

Expression of 5-aminolevulinic acid synthase in recombinant Escherichia coli

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