5(4)-Amino-4(5)-Imidazolecarboxamide, a Precursor of Purines

Shive, William; Ackermann, W. Wilbur; Gordon, Malcolm W.; Getzendaner, M. E.; Eakin, Robert E.

doi:10.1021/ja01195a521

Cited by 141 publications

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“…Our approach was to estimate the relative contributions of the histidine and purine biosynthetic pathways for synthesis of AICAR. Sulfonamides inhibit AICAR transformylase, leading to the accumulation of AICAR and the secretion of 5-amino-4-imidazole carboxamide, which can be determined from the growth medium by the method described by Bratton and Marshall (4,24,30). The amount of AICAR produced when either purine or histidine biosynthesis is repressed or disrupted will give an estimate of the relative contributions of the two pathways to AICAR production.…”

Section: Resultsmentioning

confidence: 99%

Formyltetrahydrofolate hydrolase, a regulatory enzyme that functions to balance pools of tetrahydrofolate and one-carbon tetrahydrofolate adducts in Escherichia coli

et al. 1995

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The enzyme encoded by Escherichia coli purU has been overproduced, purified, and characterized. The enzyme catalyzes the hydrolysis of 10-formyltetrahydrofolate (formyl-FH 4 ) to FH 4 and formate. Formyl-FH 4 hydrolase thus generates the formate that is used by purT-encoded 5-phosphoribosylglycinamide transformylase for step three of de novo purine nucleotide synthesis. Formyl-FH 4 hydrolase, a hexamer with 32-kDa subunits, is activated by methionine and inhibited by glycine. Heterotropic cooperativity is observed for activation by methionine in the presence of glycine and for inhibition by glycine in the presence of methionine. These results, along with previous mutant analyses, lead to the conclusion that formyl-FH 4 hydrolase is a regulatory enzyme whose main function is to balance the pools of FH 4 and C 1 -FH 4 in response to changing growth conditions. The enzyme uses methionine and glycine to sense the pools of C 1 -FH 4 and FH 4 , respectively.There are two transformylation steps in the pathway for de novo purine nucleotide synthesis. The enzymes catalyzing these reactions in Escherichia coli are purN-encoded 5Ј-phosphoribosylglycinamide (GAR) transformylase and purHencoded 5Ј-phosphoribosyl-4-carboxamide-5-aminoimidazole (AICAR) transformylase (20). Both enzymes use 10-formyltetrahydrofolate (formyl-FH 4 ) as the formyl donor. E. coli also has an alternative GAR transformylase (13, 21) that is coded for by purT (21). This enzyme enables E. coli to incorporate formate into the purine ring, even though it has no formyl-FH 4 synthetase (8). pfl-encoded pyruvate formate lyase is utilized for the production of formate under anaerobic growth conditions (23), and a gene designated purU is required for the aerobic production of formate (18). PurU encodes a protein of 280 amino acids. The sequence from residues 85 to 280 exhibits 27% amino acid identity with GAR transformylase N (purN encoded), including 10 of 11 residues that are thought to interact with formyl-FH 4 (1). This similarity as well as mutant analysis prompted the suggestion that purU encodes an enzyme that binds formyl-FH 4 and catalyzes its hydrolysis to formate and FH 4 (18).Two observations suggested that PurU-dependent formate production is regulated (18). First, growth of an E. coli purN mutant that relies on GAR transformylase T (purT encoded) and PurU for purine synthesis was strongly inhibited by glycine. This inhibition was reversed by either formate or purines. Second, methionine had a positive effect on the growth of the purN mutant, although when supplied in equimolar concentration with glycine, it did not reverse the growth inhibition. In this paper, we describe the purification and characterization of PurU and show that it is a formyl-FH 4 hydrolase whose activity is regulated by glycine and methionine, in accord with the phenotype of the purN mutant.The importance of PurU extends beyond its role in supplying formate for the GAR transformylase T reaction. It has been noted that a purU mutant required glycine supplementation to grow at...

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Section: Resultsmentioning

confidence: 99%

Formyltetrahydrofolate hydrolase, a regulatory enzyme that functions to balance pools of tetrahydrofolate and one-carbon tetrahydrofolate adducts in Escherichia coli

et al. 1995

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“…We therefore consider that our present understanding of PurU may be quite limited. Second, if formate is derived from formyl-FH4, why is the GAR transformylase step of the purine pathway less sensitive than the AICAR transformylase step to sulfonamide inhibition (21,29,35)? Previous explanations have assumed an alternative FH4-independent GAR transformylase (see reference 20, for example).…”

Section: Discussionmentioning

confidence: 99%

purU, a source of formate for purT-dependent phosphoribosyl-N-formylglycinamide synthesis

1993

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A gene designated purU has been identified and characterized. purU is adjacent to tyrT at min 27.7 on the Escherichia coli chromosome. The gene codes for a 280-amino-acid protein. The C-terminal segment of PurU from residues 84 to 280 exhibits 27% identity with 5'-phosphoribosylglycinamide (GAR) transformylase, the product of purN. Primer extension mapping and assays of lacZ in a promoter probe vector identified two promoters giving mono- and bi-cistronic purU mRNA. Neither mRNA was regulated by purines. Mutations in either of two pairs of genes are required to block synthesis of 5'-phosphoribosyl-N-formylglycinamide (FGAR) from GAR: purN purT (purT encodes an alternative formate-dependent GAR transformylase) or purN purU. On the basis of the growth of purU, purN, and purU purN mutants, it appears that PurU provides the major source of formate for the purT-dependent synthesis of FGAR.

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“…It now seems fairly certain that p-aminobenzoic acid (p-AB) and the folic acid group of factors, in the form of a coenzyme, are concerned in the biological synthesis of purines. It has been suggested (Shive et al 1947) that the coenzyme is responsible for the condensation of a 'one-carbon unit ' with 4-amino-5-imidazolecarbo~mide, or more probably a derivative of it (Shive, 1950;Woods, 1952), to produce the purine nucleus. Since p-AB and pteroylglutamic acid were present in the medium throughout the present work, the purine requirement found may have been due to an inability to complete the synthesis of this coeazyme.…”

Section: Discussionmentioning

confidence: 99%

Growth Factors for Corynebacterium diphtheriae Strain Dundee

Dalby

Holdsworth

1956

Journal of General Microbiology

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SUMIMARY:Investigations were kontinued on the nature of the growth factor requirements of Corynebacterium diphtheriae gravis, strain Dundee. Hypoxanthine, or adenine + guanine, or adenine + xanthine, were found capable of replacing concentrates of the growth factor from liver and yeast.Nucleoside and nucleotide fractions, prepared from electrophoretically-purified ribonucleic acid, were unable to replace the free purine bases. A purine extract was obtained from an active fish-liver concentrate by copper precipitation. In this extract, a large amount of hypoxanthine and a smaller amount of xanthine were found by chromatography to be the major purine constituents; traces of adenine and guanine were also detected. Chromatographic fractionation and spectrophotometric examination of the fractions confirmed the presence of hypoxanthine and xanthine, whose concentrations were estimated.The purine extract supported less growth than the original fish-liver concentrates.Growth on the purine extract could be reproduced by a mixture of hypoxanthine and xanthine at an equivalent concentration

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5(4)-Amino-4(5)-Imidazolecarboxamide, a Precursor of Purines

Cited by 141 publications

References 0 publications

Formyltetrahydrofolate hydrolase, a regulatory enzyme that functions to balance pools of tetrahydrofolate and one-carbon tetrahydrofolate adducts in Escherichia coli

Formyltetrahydrofolate hydrolase, a regulatory enzyme that functions to balance pools of tetrahydrofolate and one-carbon tetrahydrofolate adducts in Escherichia coli

purU, a source of formate for purT-dependent phosphoribosyl-N-formylglycinamide synthesis

Growth Factors for Corynebacterium diphtheriae Strain Dundee

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