Properties of an Ecto-5’-Nucleotidase of the Renal Brush Border

M, Le Hir; Angielski, Stefan; Uc, Dubach

doi:10.1159/000173064

Cited by 8 publications

(2 citation statements)

References 18 publications

(24 reference statements)

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“…The apparent Km for 5'-AMP obtained by least-square analysis of initial velocity-substrate concentration data was 0.32 mM. It is in the same range than that measured with 5'-nucleotidase of some other cells [ 1.24], but higher than that reported for the enzyme of brush border membranes [3] or for the soluble purified enzyme [4] from rat renal cortex. The activity with /j-nitrophenylphosphate.…”

Section: Discussionsupporting

confidence: 51%

“…5'-Nucleotidase (5'-ribonucleotide phosphohydrolase, EC3.1.3.5) has been character ized as an ectoenzyme, the catalytic site of which is located at the external surface of the plasma membrane, in a variety of normal and transformed cells [1,2], and particularly in the kidney [3,4], It converts its substrate, 5'-AMP. a metabolite resulting from the cata bolism of ATP.…”

Section: Introductionmentioning

confidence: 99%

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Ecto-5’-Nucleotidase of Cultured Rat Mesangial Cells

Stefanović

Savić²,

Vlahović³

et al. 1988

Kidney Blood Press Res

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Because adenosine plays a role in the regulation of glomerular filtration rate and of the release of renin, we examined the possibility of a local source for this mediator. We found that rat cultured glomerular mesangial cells converted 5’-AMP into adenosine. The properties of the enzyme involved in the reaction were those of an ecto-5’nucleotidase: (1) the products of the reaction were generated in the extracellular fluid although no 5’-nucleotidase was released by the cells into the medium; (2) identical activities were found for cultured cells in situ and sonicated cells; (3) the diazonium salt of sulfanilic acid which is a nonpenetrating reagent inhibited up to 75% of the enzyme activity. Ecto-5’-nucleotidase activity of intact cells obeyed Michaelis-Menten kinetics. Apparent K_m for 5’-AMP was 0.32 mM. 5’-UMP was a strictly competitive inhibitor. ADP exerted a very powerful inhibitory effect and behaved also as a competitive inhibitor. ATP was inhibitory both by increasing K_m and by decreasing V_max. Ecto-5’-nucleotidase was active in the absence of divalent cations. However, Mg²⁺, Ca²⁺, Co²⁺ and Mn²⁺ were stimulatory. Zn²⁺ and Cu²⁺ suppressed the activity. Concanavalin A, a plant lectin, was markedly inhibitory, suggesting that a glycoprotein moiety was necessary to express enzyme activity. Ecto-5’-nucleotidase activity was not modified during phagocytosis of serum-treated zymosan by mesangial cells. Rat cultured glomerular epithelial cells exhibited a 5’-nucleotidase activity which was 4 times lower than that of the mesangial cells in primary culture. Activity was also present in freshly isolated glomeruli and total cortex, but at lower concentrations than in mesangial cells. These findings suggest that ecto-5’-nucleotidase of mesangial cells may be the main source of adenosine within the glomeruli. Therefore, activity of this enzyme should be essential in the regulation of the glomerular microcirculation.

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Section: Discussionsupporting

confidence: 51%

Section: Introductionmentioning

confidence: 99%

Ecto-5’-Nucleotidase of Cultured Rat Mesangial Cells

Stefanović

Savić²,

Vlahović³

et al. 1988

Kidney Blood Press Res

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Extracellular cAMP: The Past and Visiting the Future in cAMP‐Enriched Extracellular Vesicles

et al. 2021

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It is recently discovered that the cyclic nucleotide, cyclic adenosine monophosphate (cAMP) can be enriched in the extracellular vesicles (EVs) isolated from endothelial cells. In the current perspective a historical context for the discovery of the extracellular cAMP is provided. The story of extracellular cAMP through investigations addressing the molecule's role in the adenosine pathway is followed, which is widespread in mammalian physiology. The adenosine pathway mediates normal physiological conditions such as renin release, phosphate transport, etc., and participates in pathological conditions such as bronchoconstriction of the airways. Furthermore, adenosine mediated biological pathways are regulated via the receptor mediated intracellular cAMP pathway in mammalian cells. It then speculates on the question of whether cAMP enriched EVs could bypass typical receptor mediated cell signaling and directly activate cAMP signaling cascade in target cells. Preliminary studies to suggest cAMP enriched EVs are provided, added to naïve endothelial cells, results in an increase in intracellular cAMP. An alternate mechanism is proposed, apart from the traditional adenosine pathway, that extracellular cAMP may exert its effects and put into perspective how it might consider circulating cAMP moving forward.

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Localization of nucleotide pyrophosphatase in the rat kidney

1986

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Hydrolysis of NAD by a nucleotide pyrophosphatase of renal membrane fractions has been reported previously. The aim of the present study was to localize this enzyme in the rat kidney. Nucleotide pyrophosphatase was assayed in glomeruli, in three parts of the proximal tubule and in four parts of the distal tubule dissected form freeze-dried sections. Nucleotide pyrophosphatase activity, expressed in mumol X min-1 X mg protein-1, ranged between 9.8 and 32.3 in the proximal tubular segments and between 1.1 and 2.7 in the distal tubular segments. It was 3.4 in the glomeruli. The enrichment of the activity during the purification of brush border vesicles was measured. A ten-fold higher specific activity was found in the brush border vesicles as compared to the renal cortical homogenates. Thus, most of the renal nucleotide pyrophosphatase appears to be localized in the luminal membrane of the proximal tubule. A permeabilization of the membrane did not increase the activity of brush border vesicles. This indicates that all catalytic sites are accessible at the outer surface of the membrane.

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Properties of an Ecto-5’-Nucleotidase of the Renal Brush Border

Cited by 8 publications

References 18 publications

Ecto-5’-Nucleotidase of Cultured Rat Mesangial Cells

Ecto-5’-Nucleotidase of Cultured Rat Mesangial Cells

Extracellular cAMP: The Past and Visiting the Future in cAMP‐Enriched Extracellular Vesicles

Localization of nucleotide pyrophosphatase in the rat kidney

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