Effect of magnesium on the properties of zinc alkaline phosphatase.

Alkaline phosphatase of Escherichia coli, isolated by procedures which do not alter its intrinsic metal content, contains 4.0 +/- 0.3 g-atoms of tightly bound zinc per mole (Kd less than 1 muM) and 1.3 +/- 0.2 g-atoms of magnesium per mole (Bosron, W.F., Kennedy, F.S., and Vallee, B.L. (1975), Biochemistry 14, 2275-2282). Importantly, the binding of magnesium is dependent both upon pH and zinc content. Hence, the failure to assign the maximal magnesium stoichiometry to enzyme isolated by conventional procedures may be considered a consequence of the conditions chosen for optimal bacterial growth and purification of the enzyme which are not the conditions for optimal binding of magnesium to alkaline phosphatase. Under the conditions employed for the present experimental studies, a maximum of six metal sites are available to bind zinc and magnesium, i.e., four for zinc and two for magnesium. Magnesium alone does not activate the apoenzyme, but it regulates the nature of the zinc-dependent restoration of catalytic activity to apophosphatase, increasing the activity of enzyme containing 2-g-atoms of zinc five-fold and that of enzyme containing 4-g-atoms of zinc 1.4-fold. Moreover, hydrogen-tritium exchange reveals the stabilizing effects of magnesium on the structural properties of phosphatase. However, neither the KM for substrate nor the phosphate binding stoichiometry and Ki are significantly altered by magnesium. Hence, magnesium, which is specificially bound to the enzyme, both stabilizes the dynamic protein structure and regulates the expression of catalytic activity by zinc in alkaline phosphatase.     

Effect of surgically induced cholestasis on the levels of hepatic zinc and metallothionein in rat liver

Early effects of experimental cholestasis on the homeostasis of zinc (Zn) and metallothionein (MT) were studied in rats which had undergone bile duct ligation for 0, 3, 6, 9, 12, 16, 20, and 24 h. Transient increases in hepatic Zn levels were observed at 9 h but returned to control values at 12 h. Serum Zn levels increased at 24 h. Cholestasis was confirmed by increased serum alkaline phosphatase (AP) activity. MT increased at 3 h and reached a maximum level at 12 h and remained elevated even at 24 h after the onset of experimental cholestasis. No hepatocellular damage was detected according to the results of alanine aminotransferase (ALT) activities in serum. These results shown that the increases in Zn observed in liver are related to bile stagnation rather than to a hepatocellular damage and that increased MT occurs concurrently with increased hepatic Zn. These observations suggest that the cellular levels of Zn in cholestasis is regulated by homeostatic mechanisms, of which one could be mediated by MT.     

Correlation between serum alkaline phosphatase activity and blood glucose levels

Fasting blood glucose levels and serum alkaline phosphatase activity of age-matched Libyan diabetic men (168) and women (168) were determined. The mean levels of blood glucose of men and women were 227 +/- 6 and 237 +/- 5 mg/dl, respectively. The respective values of serum alkaline phosphatase were 179 +/- 5 and 199 +/- 6 IU/l. The mean serum phosphatase activity of women was significantly higher (p less than 0.001) than that of their male counterparts. A statistically significant positive correlation was found between serum alkaline phosphatase and blood glucose levels of these diabetic patients (r = 0.35; p less than 0.001).     

Magnesium,zinc, and manganese in atherosclerosis of the aorta

Atomic absorption spectrometry was used in a quantitative study of zinc, magnesium, and manganese on 71 postmortal human aortas. Samples were obtained from accident victims 35-65 years of age. Fibrous plaques had higher levels of all three elements when compared to normal aortic tissue. The difference was significant for zinc (p less than 0.001) and magnesium (p less than 0.01). The high elemental levels may be both a cause and effect of atherosclerosis or the result of another unknown parameter.     

ABO blood groups, intestinal alkaline phosphatase and haptoglobin types in patients with serum hepatitis

A series of 150 patients with serum hepatitis were examined for the incidence of the Australia antigen (HBsAg) and associations with ABO blood groups, haptoglobin types and occurrence of intestinal serum alkaline phosphatase. Among the patients studied 11.3% were positive for HBsAg. When compared to controls patients with blood group O showed a significantly increased risk for serum hepatitis (p less than 0.05), while those with group B showed a decreased risk (p less than 0.01). The presence of the intestinal fraction of alkaline phosphatase showed a negative association with serum hepatitis (p less than 0.01) and there was no significant association between alkaline phosphatase types and ABO groups among the patients. The frequency of the Hp1 gene was significantly increased (p less than 0.01) among the patients as compared to controls.     

The Correlation of Serum Trace Elements and Heavy Metals with Carotid Artery Atherosclerosis in Maintenance Hemodialysis Patients

Changes in essential trace elements and heavy metals may affect the atherosclerotic state of patients on maintenance hemodialysis (HD). The aim of the study was to evaluate the relation between the serum levels of some trace elements and heavy metals (iron, zinc, manganese, copper, magnesium, cobalt, cadmium, lead, and copper/zinc ratio) and carotid artery intima-media thickness (CIMT) in HD patients. Fifty chronic HD patients without known atherosclerotic disease and 48 age- and sex-matched healthy individuals were included in the study. The serum levels of trace elements (iron, zinc, manganese, copper, and magnesium) and heavy metals (cobalt, cadmium, and lead) were measured by Atomic Adsorption Spectrophotometer (UNICAM-929). CIMT was assessed by carotid artery ultrasonography. The serum levels of iron, zinc, and manganese were lower; levels of copper, magnesium, cobalt, cadmium, lead, and copper/zinc ratio were higher in HD patients compared to controls. CIMT in HD patients were higher than the control group (0.64?±?0.11 vs 0.42?±?0.05, p?相似文献   

Effect of pH on the modulation of rat osseous plate alkaline phosphatase by metal ions.

1. Metal ions other than zinc and magnesium were effective in modulating the activity of rat osseous plate alkaline phosphatase. 2. Increasing pH had remarkable effects on the modulation of rat osseous plate alkaline phosphatase. 3. The modulation of enzyme activity by zinc, manganese and cobalt ions was slightly affected by pH variations. 4. Zinc ions were stimulatory for the enzyme at very low concentrations (50 nM). Above 50 nM zinc ions inhibited the enzyme by displacing magnesium ions. 5. Calcium ions were inhibitors of alkaline phosphatase (Kd = 10 microM) whereas manganese (Kd = 1.3 microM) and cobalt (Kd = 0.2 microM) ions were stimulatory in the pH range 8.0-10.0.     

Vitamin A toxicity and hypercalcaemia in chronic renal failure.

Serum vitamin A concentrations were measured in 38 patients undergoing haemodialysis, 24 of whom were taking multivitamin preparations containing vitamin A. Vitamin A concentrations were significantly higher in patients undergoing haemodialysis than in 28 normal controls (p less than 0.001). Patients taking vitamin A supplements had significantly higher vitamin A concentrations than those not taking them (p less than 0.05), and hypercalcaemic patients had higher concentrations than normocalcaemic patients (p less than 0.005). Withdrawal of vitamin A supplements in seven patients caused significant falls in serum vitamin A concentrations and plasma calcium concentrations (p less than 0.01 at two and three months in both cases) and in plasma alkaline phosphatase concentrations (p less than 0.01 at two months). Vitamin A toxicity can contribute to hypercalcaemia in patients undergoing haemodialysis, probably by an osteolytic effect. Multivitamin preparations containing vitamin A should therefore be prescribed with caution in these patients.     

Protective effects of zinc in chlorpyrifos induced hepatotoxicity

The toxicological literature is replete with studies attempting to explain the mechanism of action of organophosphorus (OP) insecticides to their anticholinesterase activities, but not much is known about the metabolism and detoxification of these compounds. The goal of this study was to ascertain the toxic effects of chlorpyrifos, one of the most widely used OPs, on the liver of male rats and also to evaluate the protective potential of zinc in mediating its toxic effects. It was observed that chlorpyrifos (13.5 mg/kg body weight) treatment resulted in significant inhibition (p < 0.001) of serum and hepatic acetylcholinesterase (AChE) activities after 8 wk. However, zinc-treated (227 mg/L drinking water) animals resulted in significant normalization of the inhibited AChE activities. Similarly, a significant increase in the levels of various serum and liver marker enzymes (viz. alkaline phosphatase, aspartate aminotransferase [AST], and alanine aminotransferase [ALT]) was observed following treatment with chlorpyrifos. However, coadministration of zinc to these animals restored these enzymes to within normal limits, even though some increase in the activity of serum ALT and hepatic alkaline phosphatase still persisted at the end of the study. Chlorpyrifos treatment diminished serum and hepatic zinc levels significantly (p < 0.01 to p < 0.001) compared to normal control animals. Serum iron concentrations also plummeted significantly following chlorpyrifos treatment. On the contrary, serum copper levels were significantly increased (p < 0.01) in chlorpyrifos-treated animals, but they were normalized following zinc supplementation to the rats in this group. Interestingly, chlorpyrifos treatment resulted in elevated hepatic levels of copper, iron, and selenium, but zinc treatment could only partially restore the raised elemental concentrations. These data clearly demonstrate the potential role of zinc in mediating the toxic effects of chlorpyrifos, presumably because of their antioxidant properties and also their possible interaction with other trace elements in maintaining the cellular harmony.     

Activity of the gamma-glutamyltransferase, leucine aminopeptidase and alkaline phosphatase enzymes in human tear fluid

In this work the presence of gamma-glutamyltransferase (GGT), leucine amino-peptidase (LAP) and alkaline phosphatase (AP) is shown in human tear fluid. We studied these levels according to sex, age and some eye refraction defects. The differences between the levels for both sexes are not significant. LAP and AP do not show any differences in either age groups or individuals with some refraction defects. The average level of GGT is higher from 40 years of age upwards (p less than 0.005). In individuals with refraction defects, the enzymatic activity is significantly higher (p less than 0.05) than the activities found in normal subjects. The levels of the three enzymes in serum and tear fluid do not show a significant correlation nor are they significantly modified after the samples have been frozen for a month at -20 degrees C.     

丙二醛通过激活p38和JNK通路抑制间充质干细胞成骨分化

为了探索丙二醛（malonaldehyde, MDA）抑制间充质干细胞（mesenchymal stem cells,MSCs）成骨分化的作用机制,用不同浓度的丙二醛孵育MSCs,进行成骨诱导培养,检测碱性磷酸酶活性和钙结节形成|并检测p38和JNK表达水平和磷酸化程度,用这两种信号分子的特异性阻断剂进行验证. 结果发现,丙二醛浓度依赖性地降低MSCs碱性磷酸酶的活性,抑制钙结节的形成|并引起p38和JNK的表达上调和磷酸化增强,诱导JNK由胞浆向胞核转位| p38和JNK阻断剂对丙二醛的上述效应有拮抗作用. 结果表明,丙二醛可抑制MSCs的成骨诱导分化,其作用机制涉及p38和JNK信号通路的参与.     

Histochemical study of enzyme patterns in the human submaxillary gland

Summary The histochemical distribution of various enzymes, such as alkaline phosphatase, acid phosphatase, esterase, -glycosidase, aminopeptidase, succinic dehydrogenese and TPN diaphorase, in human submaxillary glands has been determined.Acini and ducts of human submaxillary gland were devoid of alkaline phosphatase activity, but this enzyme was observed in capillaries and somewhat in myoepithelium.Activities of acid phosphatase, esterase, -glucuronidase and -galactosidase were generally observed in the entire cytoplasm of serous acini; but the cytoplasm of mucous acini was either negative or showed only trace amounts.Aminopeptidase reaction of both acini and ducts was generally negative.Succinic dehydrogenase and TPN diaphorase activities were strongly active in intralobuler ducts. Serous acini exhibited less activity with these enzymes; and mucous cells showed still less and were almost negative. In serous acini, there was much greater activity of TPN diaphorase than of succinic dehydrogenase.With 7 Figures in the Text     

The preparation of monoclonal antibodies to human bone and liver alkaline phosphatase and their use in immunoaffinity purification and in studying these enzymes when present in serum.

1. Liver and bone alkaline phosphatase isoenzymes were solubilized with the zwitterionic detergent sulphobetaine 14, and purified to homogeneity by using a monoclonal antibody previously raised against a partially-purified preparation of the liver isoenzyme. Both purified isoenzymes had a specific activity in the range 1100-1400 mumol/min per mg of protein with a subunit Mr of 80,000 determined by SDS/polyacrylamide gel electrophoresis. Butanol extraction instead of detergent solubilization, before immunoaffinity purification of the liver enzyme, resulted in the same specific activity and subunit Mr. The native Mr of the sulphobetaine 14-solubilized enzyme was consistent with the enzyme being a dimer of two identical subunits and was higher than that of the butanol-extracted enzyme, presumably due to the binding of the detergent micelle. 2. Pure bone and liver alkaline phosphatase were used to raise further antibodies to the two isoenzymes. Altogether, 27 antibody-producing cell lines were cloned from 12 mice. Several of these antibodies showed a greater than 2-fold preference for bone alkaline phosphatase in the binding assay used for screening. No antibodies showing a preference for liver alkaline phosphatase were successfully cloned. None of the antibodies showed significant cross-reaction with placental or intestinal alkaline phosphatase. Epitope analysis of the 27 antibodies using liver alkaline phosphatase as antigen gave rise to six groupings, with four antibodies unclassified. The six major epitope groups were also observed using bone alkaline phosphatase as antigen. 3. Serum from patients with cholestasis contains soluble and particulate forms of alkaline phosphatase. The soluble serum enzyme had the same size and charge as butanol-extracted liver enzyme on native polyacrylamide-gel electrophoresis. Cellulose acetate electrophoresis separated the soluble and particulate serum alkaline phosphatases as slow- and fast-moving forms respectively. In the presence of sulphobetaine 14 all the serum enzyme migrated as the slow-moving form on cellulose acetate electrophoresis. Monoclonal anti-(alkaline phosphatase) immunoadsorbents did not bind the particulate form of alkaline phosphatase in cholestatic serum but bound the soluble form. In the presence of sulphobetaine 14 all the cholestatic serum alkaline phosphatase bound to the immunoadsorbents. 4. The electrophoretic and immunological data are consistent with both particulate and soluble forms of alkaline phosphatase in cholestatic serum being derived from the hepatocyte membrane.     

Evaluation of key gluconeogenic enzymes in experimental biliary obstruction

In order to evaluate the usefulness of key gluconeogenic enzymes, in relation to the markers commonly used (alkaline phosphatase and gamma-glutamyl transpeptidase) for the diagnose of cholestasis the serum activity of phosphoenolpyruvate carboxykinase, fructose 1,6-bisphosphatase and glucose-6-phosphatase has been measured in rats with bile-duct ligation. Among the gluconeogenic enzymes studied only phosphoenolpyruvate carboxykinase activity increased significantly in the first 48 hours after cholestasis, decreasing thereafter to normal values. Both alkaline phosphatase and gamma-glutamyl transpeptidase activities showed a very significant increase which persisted throughout the experiment. These results seem to indicate that in spite of the high organ specificity of these enzymes they do not appear to be useful for the diagnosis of cholestasis.     

Prediction of early death after therapeutic hepatic arterial embolisation.

A consecutive series of 14 patients with hepatic malignant disease treated by palliative hepatic arterial embolisation was reviewed. Twelve patients had hepatic pain from their tumour and two were suffering from the carcinoid syndrome. Six patients died within four weeks of the procedure (group 1) and eight survived for at least 10 weeks (group 2). Factors were sought that might permit prediction of a high risk of early death (group 1). The pre-embolisation angiograms reviewed by a "blind" observer showed no differences in vascularity or tumour size between the groups and no difference in the extent of arterial occlusion after embolisation. The portal vein was patent in all patients. No significant difference was seen between the groups in the pre-embolisation biochemical values, with the exception of lower serum albumin concentrations and higher alkaline phosphatase activities in group 1. All those who died early had serum alkaline phosphatase activities of 45 KAU or above, while six of the eight who survived longer had activities below this value (p less than 0.02). These findings suggest that serum alkaline phosphatase activity of 45 KAU or more (normal range 3-13) might alone be a useful predictor of early death. Stepwise discriminant analysis using a weighted combination of serum alkaline phosphatase activity and albumin concentration predicted the outcome in all but one of the patients studied (p less than 0.002).     

Aldosterone and magnesium in essential arterial hypertension

Thirty mildly hypertensive patients and 27 patients with severe essential hypertension and high levels of aldosterone were selected for a study of the relationship between plasma aldosterone and magnesium in essential arterial hypertension; levels of calcium and potassium were also studied. Thirty-six individuals were used as a control group. Our findings indicate that as plasma aldosterone levels increase, serum magnesium levels decrease correspondingly: in mild hypertensives with low levels of plasma aldosterone p less than 0.05 and in the most severely hypertensive patients with high levels of plasma aldosterone p less than 0.001. In this latter group we also found an inverse correlation between serum magnesium and systolic arterial pressure (p less than 0.001) and diastolic pressure (p less than 0.01). In these patients a significant increase in urinary excretion of magnesium was found, with levels 3 times higher than in the control group. These findings suggest a close relationship between changes in plasma aldosterone and magnesium. Possibly the aldosterone contributes through this mechanism to maintaining the hypertensive state in essential arterial hypertension. This action is exercised directly through the kidney, leading to a small but constant urinary loss of magnesium. This in turn leads to a chronic depletion of magnesium in hypertensives who have high levels of plasma renin activity and highly elevated plasma aldosterone.     

Alteration of tissue zinc distribution and biochemical analysis of serum following pinealectomy in the rat

Eight weeks following pinealectomy in adult male Wistar rats, zinc levels of various tissues were found to be significantly altered: zinc in thoracic aorta was significantly increased, and in serum, pituitary, adrenal, heart, lung, and body hair, it was decreased. Serum biochemical analysis indicated that there was a significant elevation of cholesterol, alkaline phosphatase, sodium, urea, and creatinine in serum from pinealectomised rats. Liver, spleen, and thymus weights were lower following pinealectomy, although hearts were increased. The effects of pinealectomy on zinc levels in serum and tissues and on serum cholesterol and alkaline phosphatase may be related to its effects on vascular reactivity and liver fibrosis.     

Nucleotide sequence of the gene for alkaline phosphatase of Thermus caldophilus GK24 and characteristics of the deduced primary structure of the enzyme

The gene encoding Thermus caldophilus GK24 (Tca) alkaline phosphatase was cloned into Escherichia coli. The primary structure of Tca alkaline phosphatase was deduced from its nucleotide sequence. The Tca alkaline phosphatase precursor, including the signal peptide sequence, was comprised of 501 amino acid residues. Its molecular mass was determined to be 54? omitted?760 Da. On the alignment of the amino acid sequence, Tca alkaline phosphatase showed sequence homology with the microbial alkaline phosphatases, 20% identity with E. coli alkaline phosphatase and 22% Bacillus subtilis (Bsu) alkaline phosphatases. High sequence identity was observed in the regions containing the Ser-102 residue of the active site, the zinc and magnesium binding sites of E. coli alkaline phosphatase. Comparison of Tca alkaline phosphatase and E. coli alkaline phosphatase structures suggests that the reduced activity of the Tca alkaline phosphatase, in the presence of zinc, is directly involved in some of the different metal binding sites. Heat-stable Tca alkaline phosphatase activity was detected in E. coli YK537, harboring pJRAP.     

Effect of membrane moiety and magnesium ions on the inhibition of matrix-induced alkaline phosphatase by zinc ions

1. The inhibition of matrix-induced alkaline phosphatase by zinc ions is due to the displacement of magnesium ions from its binding site. 2. Binding of magnesium ions to alkaline phosphatase induces conformational changes which activate the enzyme. 3. Binding of zinc ions to alkaline phosphatase induces conformational changes which impair the catalytic action of the enzyme. 4. The inhibition of the enzyme by zinc ions is affected by membrane environment and magnesium ions.     

Response of blood serum constituents to production of and recovery from a kwashiorkor-like syndrome in the young pig.

Twenty-six 3-week-old genetically obese pigs were fed in two experiments to determine the serum chemistry profile during severe protein malnutrition and repletion. Severe protein deficiency was produced in pigs fed the high-fat, low-protein diet (growth failure, rough hair, low serum total protein and albumin). In Experiment 1, blood was sampled from the anterior vena cava of each pig five times during depletion and three times during repletion to determine serum total cholesterol, high density lipoprotein (HDL)-cholesterol, triglycerides, total protein, albumin, glucose, Ca, inorganic P, Mg, Na, K, Cl, total bilirubin, urea N, creatinine, alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, and gamma-glutamyltransferase. In Experiment 2, blood was sampled weekly for 8 weeks for serum total cholesterol, HDL-cholesterol, triglycerides, albumin, glucose, Ca, P, Mg and alkaline phosphatase. HDL-cholesterol was increased (P less than 0.01) and albumin was decreased (P less than 0.01) in protein-deficient pigs in both experiments. Creatinine, total bilirubin, gamma-glutamyltransferase, alanine aminotransferase, and aspartate aminotransferase were elevated in protein-deficient pigs compared with controls after 7 weeks of depletion. Inorganic P (P less than 0.01), Ca (P less than 0.01), and Mg (P less than 0.05) concentrations were depressed in protein-depleted pigs compared with controls in both experiments. After 8 weeks of repletion in Experiment 1, all elements except inorganic P were similar in the two groups. Short-term, severe, protein malnutrition affected lipid, electrolyte, and structural mineral metabolism and indices of liver function in the absence of parasites, diarrhea, and infection. The effects were reversed after 8 weeks of repletion. We conclude that the elevated serum cholesterol in protein deficiency is related primarily to an increase in the HDL fraction.