The formation of 5-formyluracil by cell-free preparations from Neurospora crassa

Abbott, Mitchel T.; Dragila, T.A.; McCroskey, R.P.

doi:10.1016/0005-2787(68)90002-6

Cited by 30 publications

(7 citation statements)

References 11 publications

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“…2-Keto-1-14C1 -glutaric acid was obtained from New England Nuclear Corp., Frankfurt/Main, Germany. 5-Formyluracil was synthesized as described previously [9] . IV.…”

Section: Methodsmentioning

confidence: 99%

“…The mold was grown in minimal medium [7] . Mycelia were harvested and homogenized as previously reported [9]. After dialysis of the 100,OOOg supernatant fraction against 10 mM potassium phos- and thymidine 2'-hydroxylase (e-e-e) on a column (25 X 125 mm) of DEAESephadex (A-50).…”

Section: Methodsmentioning

confidence: 99%

“…Assays were based on the finding that 2-ketoglutarate is.oxidatively decarboxylated concomitantly with oxygenation of thymine [7] ,.5hydroxymethyl-uracil, 5-formyluracil [9] , 1-methyluraciI, thymidine, and deoxyuridine (unpublished) The incubation mixture (0.2 ml) had the following composition: Xl-300 mg enzyme per 1,0.5 mM pyrimidine or nucleoside substrate, 0.25 mM potassium 2-keto-l-r4C1-glutarate (0.25 mCi per l), 5 mM FeS04, 5 mM sodium ascorbate, and 50 mM potassium phosphate at pH 7.5. Incubations were carried out for 30 min at 37' and then terminated by addition of trichloracetic acid (50 g/l).…”

Section: Methodsmentioning

confidence: 99%

“…Isoelectric focusing showed a pI value of 4.6 for thymidine hydroxylase and of 4.9 for thymine 7-hydroxylase. Thymidine 2'-hydroxylase has similar cofactor requirements as thymine 7hydroxylase, indicating a similar reaction mechanism [9] . The biological role of thymidine 2'-hydroxylase is unknown at present.…”

Section: Hydroxylation Of Thymidinementioning

confidence: 98%

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Oxygenases involved in thymine and thymidine metabolism inNeurospora crassa

Bankel¹,

Holme²,

Lindstedt³

et al. 1972

FEBS Letters

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“…2-Keto-1-14C1 -glutaric acid was obtained from New England Nuclear Corp., Frankfurt/Main, Germany. 5-Formyluracil was synthesized as described previously [9] . IV.…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Hydroxylation Of Thymidinementioning

confidence: 98%

See 2 more Smart Citations

Oxygenases involved in thymine and thymidine metabolism inNeurospora crassa

Bankel¹,

Holme²,

Lindstedt³

et al. 1972

FEBS Letters

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“…The specific requirement for ferrous ion and 2-ketoglutarate is found also for rat liver y-butyrobetaine hydroxylase (Lindstedt, 1967a-c), collagen proline hydroxylase (Hutton et al, 1966(Hutton et al, , 1967Kivirikko and Prockop, 1967a,b), collagen lysine hydroxylase (Hausmann, 1967), and for thymine 7-hydroxylase and 5-hydroxymethyluracil7-oxygenase from Neurospora (Abbott et al, 1967(Abbott et al, , 1968. A stoichiometry between 2-ketoglutarate degradation and hydroxylation occurs also for the hydroxylation of collagen proline (Rhoads and Udenfriend, 1968) and for the hydroxylation of thymine to 5-hydroxymethyluracil (Holme et a/., 1970).…”

Section: -B U T Y R O B E T a I N E H Y D R O X Y L A S Ementioning

confidence: 96%

γ-Butyrobetaine hydroxylase from Pseudomonas sp AK 1

Lindstedt¹,

Lindstedt²,

Tofft³

1970

Biochemistry

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A soluble y-butyrobetaine hydroxylase has been partially purified from cells of a Pseudomonas strain (Pseudomonas sp AK 1) by chromatography on DEAE-cellulose and on hydroxylapatite. Carnitine is the only product of y-butyrobetaine; trimethylamine or trimethylaminoacetone is not formed. The enzyme has a low isoelectric point (around pH 4.5). The K M ,~~~ value for y-butyrobetaine is 2.4 mM. There is an absolute and specific requirement for 2-ketoglutarate and I n the previous studies of carnitine metabolism, evidence was obtained for a low turnover rate of carnitine in the rat (Lindstedt and Lindstedt, 1961a). Of various hypothetical precursors, only y-butyrobetaine (4-trimethylaminobutyrate) was converted efficiently into carnitine Lindstedt, 1961 b, 1965a). Results from studies with preparations from rat liver indicated an oxygenase type of mechanism for this reaction (Lindstedt and Lindstedt, 1962;. The pathway for carnitine degradation in the rat is unknown. A poxidation type of reaction for the degradation of carnitine in Pseudomonas aeruginosa NCTC A 7244 was suggested from the finding of glycine betaine as a metabolite of carnitine (Lindstedt and Lindstedt, 1962). The possibility that this type of mechanism occurred in the formation of carnitine from ybutyrobetaine in bacteria was therefore considered. A Pseudomonas strain (Pseudomonas sp AK 1) was obtained by enrichment culture on y-butyrobetaine media (Lindstedt et af., 1967b). Carnitine was formed from y-butyrobetaine in incubations with acetone-dried cells under conditions which suggested a monooxygenase reaction. The hydroxylase from Pseudomonas sp AK 1 has been used for studies of the reaction mechanism (Lindblad et al., 1969) as high specific activity is more easily obtained with the bacterial preparations than with those from rat liver. The partial purification of the enzyme and studies of cofactor requirements are now presented. Some of the results have been reported in preliminary form (Lindstedt et af., 1967a,c). Material and MethodsChemicals. Chemicals were obtained from the following sources: y-butyrobetaine chloride, ferrous sulfate, and 1,lOphenanthroline from E. Merck, AG, Darmstadt, West Germany; oxalacetic acid, mercaptoethanol, L-glutamic acid, sodium DL-isocitrate, zinc 2-hydroxyglutarate, sodium pyruvate, succinic acid, isoascorbate, dichlorophenolindophenol, and

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