Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Renal brush-border membrane vesicles from rat kidney cortex were irradiated in frozen state with a gamma-radiation source. Initial rates of influx into these vesicles were estimated for substrates such as L-glutamic acid, L-alanine, L-proline and L-leucine to establish the molecular sizes of their carriers. Transport was measured in initial-rate conditions to avoid artifacts arising from a decrease in the driving force caused by a modification of membrane permeability. Initial rates of Na(+)-independent uptakes for those four substrates appeared unaffected in the dose range used (0-6 Mrad), indicating that the passive permeability of the membrane towards these substrates was unaffected. However, at higher doses of irradiation the Na+ influx and the intravesicular volume evaluated by the uptake of glucose at equilibrium were altered by radiation. Thus Na(+)-dependent influx values were corrected for volume changes, and the corrected values were used to compute radiation-inactivation sizes of the transport systems. Their respective values for L-glutamic acid, L-proline, L-leucine and L-alanine carriers were 250, 224, 293 and 274 kDa. The presence of the free-radicals scavenger benzoic acid in the frozen samples during irradiation did not affect the uptake of glucose, phosphate and alkaline phosphatase activity. These results indicate that freezing samples in a cryoprotective medium was enough to prevent secondary inactivation of transporters by free radicals. Uptakes of beta-alanine and L-lysine were much less affected by radiation. The radiation-inactivation size of the Na(+)-dependent beta-alanine carrier was 127 kDa and that of the L-lysine carrier was 90 kDa.
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PMID:Molecular sizes of amino acid transporters in the luminal membrane from the kidney cortex, estimated by the radiation-inactivation method. 197 9

High levels of interleukin-6 (IL-6) have been detected in synovial fluid from patients with inflammatory arthropathies associated with local bone resorption, suggesting a role for IL-6 as a local regulator of bone resorption and remodeling. In the present study we examined the effects of IL-6 on [3H]thymidine ([3H]TdR) incorporation, collagen synthesis, and alkaline phosphatase activity in UMR-106-01 rat osteoblastic osteosarcoma cells. IL-6 stimulated a dose-dependent increase in [3H]TdR incorporation that was maximal at 1000 U/ml (-147% of basal, p less than 0.005) in osteoblastlike cells that were in a logarithmic phase of growth. The increase in [3H]TdR incorporation was maximal between 12 and 24 h and was neutralized by pretreatment with the polyclonal rabbit antibody to IL-6. IL-6 also increased cell number and the secretion of prostaglandin E2 in UMR-106-01 cells in logarithmic growth phase. The stimulation of [3H]TdR incorporation and release of PGE2 into the culture medium by IL-6 was inhibited by indomethacin. A 24 h exposure of the osteoblastlike cells to 1000 U/ml of IL-6 reduced [3H]proline incorporation into collagenase-digestible (CDP) protein to 73% of control values (p less than 0.01). Noncollagen protein (NCP) synthesis was inhibited to 80% of control values (p less than 0.01) by 1000 U/ml of IL-6. The inhibitory effect was relatively greater on CDP than on NCP and consequently resulted in a decrease in the percentage of collagen synthesis. Alkaline phosphatase activity was not altered in these cells after a 24 h exposure to 1-1000 U/ml of IL-6.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effects of interleukin-6 on cellular function in UMR-106-01 osteoblastlike cells. 202 35

Calcium channel blockers have been reported to have such diverse effects on reduction in protein synthesis, diminished incorporation of proline into new collagen, and decreased hormone release in vitro. The chronic affect of the calcium channel blocker nifedipine was examined in vivo to determine the possible impact of pharmacologic calcium channel blockade on bone metabolism. Eleven Caucasian males treated with an average of 40 mg/d nifedipine for an average of three years were compared to 11 control males matched for age, height, weight, activity level, cardiovascular status, and calcium intake. No significant differences between groups were noted in bone mineral density at the lumbar spine (L2-4), proximal femur (femoral neck, Ward's triangle and trochanter), and proximal and distal radius. There were also no significant differences in parameters of bone turnover (alkaline phosphatase, osteocalcin, urine calcium/creatinine, and hydroxyproline/creatinine ratio), or hormones that might affect calcium metabolism and bone (testosterone, PTH, 25(OH) vitamin D, and calcitonin). In summary, chronic nifedipine use in males is not associated with either a beneficial or adverse effect on bone metabolism.
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PMID:Chronic use of the calcium channel blocker nifedipine has no significant effect on bone metabolism in men. 205 35

The present study examined the effect of long-term, moderate physical exercise on trabecular bone volume (TBV), calcium content, 3H-proline uptake, and the activities of alkaline and acid phosphatases in lumbar vertebrae of aging and senescent mice. It became apparent that if physical activity starts at an early stage of life, i.e., prior to middle age and is extended until old age, it exerts beneficial effects on trabecular bone mass and mineralization. Such a positive effect is not obtained if the training program is initiated after middle age. The training-induced reduction in bone loss was accompanied by a significant decrease in acid phosphatase activity whereas no changes took place with regard to the activity of alkaline phosphatase. Long-term physical exercise also enhanced the uptake of 3H-proline by lining cells along the bone trabecules. In spite of its moderate nature, the endured training program served as a stress factor for the involved animals, a fact that was manifested by an increase in the serum levels of corticosterone. Thus, it seems that whereas young animals respond favorably to such a stimulatory stress, older animals lose this ability of adaptation.
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PMID:Long-term physical exercise retards trabecular bone loss in lumbar vertebrae of aging female mice. 210 53

Recombinant human interferon-alpha 2C and recombinant human interferon-gamma (5-1000 U/ml) inhibit the proliferation of normal human bone-derived cells and a human osteosarcoma cell line. In the bone-derived cells the inhibitory effect of interferon-gamma was significantly greater than that of interferon-alpha, whereas in the osteosarcoma cell line the inhibitory effects of both interferons were quantitatively similar. Interferon-alpha did not affect the alkaline phosphatase activity of either type of cells. In contrast, interferon-gamma affected the activity of the enzyme in both cell types: in the bone-derived cells the effect of interferon-gamma was stimulatory whereas in the osteosarcoma cells the effect was inhibitory. In both cell types interferon-gamma selectively inhibited the incorporation of radiolabelled proline into type I collagen. In the osteosarcoma cells, the effects of both interferons on collagen synthesis were quantitatively similar. In the bone-derived cells, however, interferon-alpha decreased proline incorporation into collagen and non-collagen proteins to a similar extent and thus did not affect collagen synthesis when expressed as a percentage of total protein synthesis. Two-dimensional polyacrylamide gel electrophoresis of the radiolabelled proteins of the cell layer synthesised by both cell types in the presence of either interferon demonstrated that this treatment enhanced or induced the synthesis of a total of 21 individual proteins (19 in bone cells, 14 in osteosarcoma), ranging in apparent molecular mass over 14-87 kDa. The set of proteins induced was different in all four combinations of cells and interferon. A tentative identification of several of the proteins was possible based upon estimation of molecular mass, preferential induction by interferon-alpha or interferon-gamma and differential induction in normal and transformed bone-derived cells. The results of this study demonstrate that interferons have complex effects upon the proliferative and biosynthetic activities of human bone-derived cells and demonstrate significant differences between the responses of normal cells and transformed bone-derived cell line. Further investigations will be required in order to determine whether or not these differences are unique to the osteosarcoma cell line or are a characteristic of the effects of interferons on bone-derived cells in general.
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PMID:Interferons and bone. A comparison of the effects of interferon-alpha and interferon-gamma in cultures of human bone-derived cells and an osteosarcoma cell line. 212 83

Effects of transforming growth factor (TGF)-beta, epidermal growth factor (EGF), insulin, 1, 25-dihydroxyvitamin D3 (1, 25 (OH)2D3), and parathyroid hormone (PTH) on the proliferation and differentiation of clonal dental pulp cells of rats were investigated. Interaction between growth factors (TGF-beta and EGF) and two hormones insulin and 1, 25 (OH)2D3, which have been noticed to accelerate the differentiation of the cells, were also studied, and the following results were obtained: 1) TGF-beta decreased alkaline phosphatase (ALPase) activity in a dose-dependent manner, and the inhibitory effect was not blocked by indomethacin, suggesting that the effect of TGF-beta on the cells may not be mediated by prostaglandins. Inhibitory effects of ALPase antagonists (L-phenylalanine, L-homoarginine, levamisole) on the activity were not affected by TGF-beta. TGF-beta showed no evident effect on the DNA synthesis (incorporation of [3H] thymidine) and collagen synthesis (incorporation of [2, 3-3H] proline into the collagenase-digestible protein) of the cells. 2) EGF stimulated the incorporation of [3H] thymidine and inhibited the ALPase activity. The inhibitory effect was not blocked by indomethacin, indicating that the EGF effect is not mediated by prostaglandins. Collagen synthesis was significantly inhibited by EGF. 3) Insulin showed a weak but significant inhibition of the DNA synthesis. Insulin increased the ALPase activity evidently, and accelerated the collagen synthesis significantly. 4) The vitamin 1, 25 (OH)2D3 significantly increased the ALPase activity though no significant changes were observed in the DNA synthesis and collagen synthesis. 5) PTH had no evident effect on the DNA synthesis and ALPase activity, but did tend to accelerate the collagen synthesis. 6) A study on the interaction between insulin and EGF or TGF-beta revealed that the acceleration of DNA synthesis induced by EGF was inhibited when the factor was combined with insulin, and the increase in ALPase activity elicited by insulin was inhibited by EGF and weakened by TGF-beta significantly when these factors were added simultaneously with the insulin. Or viewed another way, the inhibitory effect of EGF or TGF-beta on the ALPase activity was antagonized by insulin. The accelerative action of insulin on collagen synthesis was antagonized by EGF and potentiated by TGF-beta. 7) A study on the interaction between 1, 25 (OH)2D3 and EGF or TGF-beta revealed that 1, 25 (OH)2D3 inhibited the accelerating effect of EGF on the DNA synthesis and that the increasing effect of 1, 25 (OH)2D3 on ALPase activity was strongly inhibited by EGF.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:[Effects of various growth factors and hormones on clonal rat pulp cells]. 213 79

The predicted amino acid sequence of the alpha subunit of the rat liver mitochondrial ATP synthase has been obtained by sequencing a cDNA for the alpha subunit. Analysis of the sequence shows that it contains the A and B consensus sequences found in many nucleotide-binding proteins. Twelve amino acids of the rat liver alpha subunit differ from the sequence of the bovine heart alpha subunit; four of these involve differences in charge. The rat liver alpha subunit, from arginine 15 to the C-terminal proline 510, has been overexpressed in Escherichia coli using the alkaline phosphatase promoter (phoA) and leader peptide to direct the export of the expressed protein to the bacterial periplasm. By treating the cells with lysozyme, osmotic shock, and alkaline pH washes, the alpha subunit can be extracted in high yield (greater than 25 mg/liter) and in a high state of purity. The expressed alpha subunit remains soluble at pH 9.5 or greater and precipitates when treated with Mg2+ ions at low millimolar concentration. The bacterially expressed alpha subunit interacts with 2'(3')-O-(2,4,6-trinitrophenyl)adenosine 5'-triphosphate (TNP-ATP), resulting in a marked fluorescence enhancement upon binding. An enhancement of fluorescence is also observed upon the interaction of the alpha subunit with TNP-ADP. Preincubating the alpha subunit with 1.5 mM ATP significantly reduces the fluorescence enhancement seen with TNP-ATP. The alpha subunit binds TNP-ATP with an apparent Kd in the low micromolar range (1-5 microM) and binds TNP-ADP with an affinity at least 10-fold lower. This work shows that the rat liver alpha subunit can be overexpressed in E. coli to yield a large amount of functional protein. With the acquisition of the overexpressed alpha subunit, it is now possible to test the reconstitution of ATPase activity from a mixture of recombinant and rat liver-derived subunits and to test the formation of complexes by the overexpressed alpha and beta subunits of the rat liver F1-ATPase.
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PMID:Mitochondrial ATP synthase. cDNA cloning, amino acid sequence, overexpression, and properties of the rat liver alpha subunit. 213 25

Many proteins are now known to be anchored to the plasma membrane by a phosphatidylinositol-glycan (PI-G) moiety that is attached to their COOH termini. Placental alkaline phosphatase (PLAP) has been used as a model for investigating mechanisms involved in the COOH-terminal processing of PI-G-tailed proteins. The COOH-terminal domain of pre-pro-PLAP provides a signal for processing during which a largely hydrophobic 29-residue COOH-terminal peptide is removed, and the PI-G moiety is added to the newly exposed Asp-484 terminus. This cleavage/attachment site was subjected to an almost saturation mutagenesis, and the enzymatic activities, COOH-terminal processing, and cellular localizations of the various mutant PLAP forms were determined. Substitution of Asp-484 by glycine, alanine, cysteine, asparagine, or serine (category I) resulted in PI-G-tailed and enzymatically active proteins. However, not all category I mutant proteins were PI-G tailed to the same extent. Pre-pro-PLAP with other substituents at position 484 (threonine, proline, methionine, valine, leucine, tyrosine, tryptophan, lysine, glutamic acid, and glutamine; category II) were expressed, as well as the category I amino acids, but there was little or no processing to the PI-G-tailed form, and this latter group exhibited very low enzyme activity. The bulk of the PLAP protein produced by category II mutants and some produced by category I mutants were sequestered within the cell, apparently in the endoplasmic reticulum (ER). Most likely, certain amino acids at residue 484 are preferred because they yield better substrates for the putative "transamidating" enzyme. In transfected COS cells, at least, posttranslational PI-G-tail processing does not go to completion even for preferred substrates. Apparently PI-G tailing is a requisite for transport from the ER and for PLAP enzyme activity. Proteins that are not transamidated are apparently retained in the ER in an inactive conformation.
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PMID:Selectivity of the cleavage/attachment site of phosphatidylinositol-glycan-anchored membrane proteins determined by site-specific mutagenesis at Asp-484 of placental alkaline phosphatase. 215 84

The purpose of the study was to investigate the effect of skeletal growth factor/insulinlike growth factor II and other growth factors known to be present in bone matrix on the proliferation and differentiation of human bone cells. Cells were isolated by collagenase digestion from femoral heads obtained during hip replacement operations. Cells were cultured in DMEM medium with 10% calf serum. Third to fifth passage cells were plated in multiwell plates and the medium changed to low serum (0.1%) for 2 days. The medium was changed to serum-free medium prior to addition of growth factors. Cell proliferation was measured by the incorporation of [3H]thymidine into DNA and by the percentage of cells that incorporate bromodeoxyuridine. Protein synthesis was measured by the incorporation of [3H]proline into trichloroacetic acid-precipitable material. Skeletal growth factor/insulinlike growth factor II and insulinlike growth factor I stimulated cell proliferation and protein synthesis in a dose-dependent manner. Alkaline phosphatase-specific activity was not increased by these factors. Transforming growth factor beta 1 did not affect cell proliferation but stimulated protein synthesis and increased the specific activity of alkaline phosphatase. Fibroblast growth factor did not affect any of the cell parameters. These studies suggest that skeletal growth factor/insulinlike growth factor II, insulinlike growth factor I, and transforming growth factor beta 1 may play a role in the local control of the proliferation and differentiation of human osteoblasts.
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PMID:Skeletal growth factor and other growth factors known to be present in bone matrix stimulate proliferation and protein synthesis in human bone cells. 215 9

Gentamicin nephrotoxicity is associated with impairments in proximal tubular function. This study determined whether gentamicin administration to the rat, before a reduction in glomerular filtration rate (GFR), causes early and selective alterations in renal cortical brush-border membrane (BBM) enzyme and transport activity, lipid composition, and fluidity. Three days of gentamicin administration caused significant decreases in the Vmax of alkaline phosphatase, the Vmax of sodium gradient-dependent phosphate transport (Na-Pi cotransport), and the Vmax of pH gradient-dependent sodium transport (Na-H exchange). Gentamicin did not affect BBM-bound maltase or leucine aminopeptidase activities and sodium gradient-dependent glucose or proline transport activities. Gentamicin also caused a significant decrease in BBM sphingomyelin, significant increases in BBM phosphatidylcholine and phosphatidylinositol, a significant decrease in the phospholipid fatty acid saturation index, and a significant increase in BBM fluidity, i.e., decrease in the fluorescence anisotropy of diphenylhexatriene. These BBM functional and compositional effects of gentamicin were independent of endogenous parathyroid hormone activity. We conclude that gentamicin causes early and specific alterations in BBM enzyme and transport activity and also lipid composition, which may play an important role in the progression of renal cell injury.
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PMID:Early selective effects of gentamicin on renal brush-border membrane Na-Pi cotransport and Na-H exchange. 215 23


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