Kinetic studies of mutant human erythrocyte adenine phosphoribosyltransferases

Henderson, J. Frank; Miller, Helen R.; Kelley, William N.; Rosenbloom, Frederick M.; Seegmiller, J. Edwin

doi:10.1139/o68-108

Cited by 25 publications

(5 citation statements)

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“…We measured PRPP availability using [ 14 C]adenine because the K m of adenine phosphoribosyltransferase for PRPP is about 1 ⁄ 25 that of the K m of hypoxanthine phosphoribosyltransferase for PRPP (38,39). Thus, assessing adenine incorporation into purine nucleotides should be more sensitive to changes in the intracellular PRPP concentration than measuring hypoxanthine or guanine incorporation into nucleotides.…”

Section: Discussionmentioning

confidence: 99%

The Phosphatidylinositol 3-Kinase/Akt Cassette Regulates Purine Nucleotide Synthesis

Wang

Fridman

Blackledge

et al. 2009

Journal of Biological Chemistry

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The phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway is highly conserved throughout evolution and regulates cell size and survival and cell cycle progression. It regulates the latter by stimulating procession through G 1 and the G 1 /S phase transition. Entry into S phase requires an abundant supply of purine nucleotides, but the effect of the PI3K/Akt pathway on purine synthesis has not been studied. We now show that the PI3K/Akt cassette regulates both de novo and salvage purine nucleotide synthesis in insulin-responsive mouse mesenchymal cells. We found that serum and insulin stimulated de novo purine synthesis in serum-starved cells largely through PI3K/Akt signaling, and pharmacologic and genetic inhibition of PI3K/Akt reduced de novo synthesis by 75% in logarithmically growing cells. PI3K/ Akt regulated early steps of de novo synthesis by modulating phosphoribosylpyrophosphate production by the non-oxidative pentose phosphate pathway and late steps by modulating activity of the bifunctional enzyme aminoimidazole-carboxamide ribonucleotide transformylase IMP cyclohydrolase, an enzyme not previously known to be regulated. The effects of PI3K/Akt on purine nucleotide salvage were likely through regulating phosphoribosylpyrophosphate availability. These studies define a new mechanism whereby the PI3K/Akt cassette functions as a master regulator of cellular metabolism and a key player in oncogenesis.Insulin and a variety of other growth factors activate the PI3K 3 /Akt (protein kinase B) pathway. Activated Akt regulates several intracellular processes including protein synthesis, glucose metabolism, and cell cycle progression (1, 2). Frequently regulation occurs at more than one step. (i) Akt increases protein synthesis by activating mTOR, which regulates the activities of S6 kinase-1 and 4E-BP1, two translational regulators (3).(ii) Akt regulates glucose metabolism by inducing translocation of glucose transporters to the cell surface, by activating hexokinase, and by inhibiting glycogen synthase kinase-3 (4). (iii) Akt stimulates the cell cycle by phosphorylating, and thereby inhibiting, the cyclin-dependent kinase inhibitors p21 Cip/WAF1 and p27Kip1 and the FOXO transcription factors and through phosphorylation of glycogen synthase kinase-3, which regulates the G 1 cyclins, cyclins D and E (2, 5).The de novo synthesis of purines consists of 10 sequential steps, starting with phosphoribosylpyrophosphate (PRPP) and ending with IMP; the latter is converted to AMP or GMP (see Fig. 1). PRPP amidotransferase catalyzes the first committed step of the pathway (see Fig. 1, reaction 2) and is subject to feedback inhibition by purine nucleotides; therefore, it has been considered a major point of pathway regulation (6). We showed previously that production of ribose 5-phosphate, the immediate precursor of PRPP and an end product of the pentose phosphate pathway, also contributes to the regulation of purine synthesis (7). Salvage purine nucleotide synthesis involves hypoxanthine phosphoribosyltransferase, which...

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Section: Discussionmentioning

confidence: 99%

The Phosphatidylinositol 3-Kinase/Akt Cassette Regulates Purine Nucleotide Synthesis

Wang

Fridman

Blackledge

et al. 2009

Journal of Biological Chemistry

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“…First, this appar¬ ently represents a significant species difference since the value obtained with the human erythrocyte enzyme is nearly ten times higher than that reported for the yeast enzyme.33 In addition, this represents a value ap¬ proximately 30 times higher than that obtained for the human adenine phosphoribosyltransferase which pre¬ sumably would allow the latter en¬ zyme to compete much more effectively for PRPP in the cell. 34 Finally, the normal concentration of PRPP in cultured human fibroblasts35 and in human erythrocytes3638 is probably less than 2% of this value. From these in vitro considerations alone, one might question the physiologic impor¬ tance of this enzyme.…”

Section: Properties Of the Normal Human Hypoxanthine-guanine Phosphormentioning

confidence: 99%

Biochemistry of the X-Linked Uric Aciduria—Enzyme Defect and Its Genetic Variants

Kelley¹

1972

Arch Intern Med

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“…The concentration of PRPP was 1 mM, except in the kinetic studies. Kinetic studies were done essentially according to the method of Henderson et al [14]. AMP and adenine were separated by high-voltage electrophoresis and the radioactivities were measured by liquid-scintilla tion counter.…”

Section: Enzymatic Assay and Kinetic Studiesmentioning

confidence: 99%

Partial and Complete Adenine Phosphoribosyltransferase Deficiency Associated with 2,8-Dihydroxyadenine Urolithiasis: Kinetic and Immunochemical Properties of APRT

Abe¹,

Hayasaka²,

Narisawa³

et al. 1987

Enzyme

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We have studied adenine phosphoribosyltransferase (APRT) in the hemolysates from the families of 2,8-dihydroxyadenine urolithiasis associated with partial deficiency of APRT (the Japanese type) and complete deficiency of APRT (the null type). The APRT in the control subjects was found to be heat-stable at the physiological concentration of phosphoribosylpyrophosphate (PRPP), which was close to the value of its Km for PRPP. The APRT in the Japanese type showed 10 times higher Km values for PRPP and needed a comparably increased level of PRPP for stability in vitro. No change in red cell PRPP was found in the Japanese type of APRT deficiency. The content of APRT enzyme protein was decreased in the hemolysates of the Japanese type, probably due to its lability at the level of PRPP present in the cells. The heterozygote of the null type also had labile enzyme molecules at the physiological PRPP concentration.

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Kinetic studies of mutant human erythrocyte adenine phosphoribosyltransferases

Cited by 25 publications

References 0 publications

The Phosphatidylinositol 3-Kinase/Akt Cassette Regulates Purine Nucleotide Synthesis

The Phosphatidylinositol 3-Kinase/Akt Cassette Regulates Purine Nucleotide Synthesis

Biochemistry of the X-Linked Uric Aciduria—Enzyme Defect and Its Genetic Variants

Partial and Complete Adenine Phosphoribosyltransferase Deficiency Associated with 2,8-Dihydroxyadenine Urolithiasis: Kinetic and Immunochemical Properties of APRT

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