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)

A new affinity label for ribulose bisphosphate carboxylase/oxygenase from Rhodospirillum rubrum, 2-(4-bromoacetamido)anilino-2-deoxypentitol 1,5-bisphosphate, has been prepared, Reductive amination of ribulose-P2 with p-phenylenediamine in the presence of sodium cyanoborohydride yielded an epimeric mixture which was resolved by chromatography on quaternary aminoethyl-Sephadex. Subsequent bromoacetylation of the isolated amino bisphosphates gave reagents A and B (ribo and arabino epimers of 2-(4-bromoacetamido) anilino-2-deoxypentitol 1,5-bisphosphate) which were competitive inhibitors of the carboxylase with Ki values of 705 and 104 microM, respectively. Reagent A exhibited no time-dependent effects on the carboxylase in either the deactivated or activated state. Incubation of the enzyme with reagent B in the presence of the essential activators CO2 and Mg2+, however, resulted in an irreversible, time-dependent loss of activity, with a Kinact of 125 microM and a minimal half-time of 7.3 min. Covalent incorporation of [14C]reagent B was directly proportional to the loss of activity, with total inactivation correlating with an incorporation of 1.1 mol of reagent/mol of subunit. Inclusion of the competitive inhibitor 2-carboxyribitol 1,5-bisphosphate protected against inactivation with a concomitant reduction in incorporation. Neither reagent affected the activity of spinach carboxylase. Fractionation of [14C]reagent B-modified enzyme on DEAE-cellulose, subsequent to carboxymethylation and tryptic digestion, revealed two major radioactive peaks of approximately equal area. Digestion of each peak with alkaline phosphatase and rechromatography on DEAE-cellulose resulted in pure peptides I and II. The peptides were identical except in the site of labeling: peptide I contained a modified cysteinyl residue while peptide II contained a modified histidyl residue. Automated Edman degradation established the sequence as (sequence in text) which is located near the NH2 terminus of the enzyme. The lack of reactivity with the spinach enzyme is explained by the deletion of the histidyl residue and the replacement of cysteine by tryptophan in the eukaryotic species. Although the nonconservation of the modified residues argues against a functional role other than maintenance of structural integrity, the extensive homology in this region among seven different species of carboxylase is compatible with the region comprising a portion of the active site.
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PMID:2-(4-Bromoacetamido)anilino-2-deoxypentitol 1,5-bisphosphate, a new affinity label for ribulose bisphosphate carboxylase/oxygenase from Rhodospirillum rubrum. Determination of reaction parameters and characterization of an active site peptide. 642 17

Two major forms of human alpha-L-iduronidase have been individually purified over 175,000-fold to apparent homogeneity by sequential anion exchange, lectin affinity, and gel filtration chromatography. The two forms, initially designated as soluble and membrane-associated, were extracted from human lung in approximately equal amounts. Optimal solubilization of the membrane-associated form was facilitated by use of a non-ionic detergent or mannose 6-phosphate and saponin. Following detergent homogenization, the two forms were separated by anion exchange chromatography and then individually purified. The more electronegative form was membrane-associated, had a pI of approximately 5.9, and was selectively taken up (high uptake) by cultured Hurler syndrome fibroblasts; the more electropositive soluble form had a pI of about 6.6 and was incorporated into Hurler fibroblasts at a markedly lower rate (low uptake). After treatment with alkaline phosphatase, the pI values of both enzymes were about 7.8. Using 4-methylumbelliferyl-alpha-L-iduronide as substrate, the low and high uptake forms were each purified in milligram quantities to specific activities of 284,000 and 202,000 units/mg, respectively, with a combined yield greater than 35%. Each purified enzyme form migrated as a single protein band which also stained for enzymatic activity when electrophoresed in 7% native polyacrylamide disc gels at pH 4.3. By gel filtration, the high uptake form had an Mr = 85,000 whereas the Mr for the low uptake form was 68,000. Molecular weight estimates by analytical polyacrylamide gel electrophoresis were 82,000 and 70,000 for the high and low uptake forms, respectively. Rabbit anti-human low uptake alpha-L-iduronidase antibodies cross-reacted with the high uptake form as demonstrated by both immunotitration and Ouchterlony double immunodiffusion. Amino acid analysis revealed that the high uptake (higher molecular weight) form contained more arginine, glycine, alanine, glutamate or glutamine, leucine, isoleucine, histidine, and proline residues per molecule than the low uptake (lower molecular weight) form. Automated Edman degradation determined that the NH2-terminal residues of both forms were blocked. Both sodium dodecyl sulfate-polyacrylamide gel electrophoresis and high performance liquid chromatography demonstrated that each purified form was composed of several components; each post-high performance liquid chromatographic component retained catalytic activity and was immunologically cross-reactive with antibodies against the low uptake form.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Human alpha-L-iduronidase. I. Purification and properties of the high uptake (higher molecular weight) and the low uptake (processed) forms. 642 18

Ten patients with protracted diarrhea of infancy received either 8% or 16% of 130 calories/kg/day as amino acids. Patients were treated initially with total parenteral nutrition (TPN) and subsequently with an elemental diet. Nitrogen balance was better during TPN in those who received the higher amino acid intake. No differences were noted between groups in weight gain or restoration of muscle mass. Higher levels of serum urea nitrogen and alkaline phosphatase were noted in patients with the higher amino acid intake; cholestatic liver injury developed in two of these patients. The patients receiving the higher amount of amino acid demonstrated enhanced calciuria during TPN. Other than better nitrogen balance, no clinical benefits and more undesirable side effects were observed in patients receiving 16% amino acid calories.
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PMID:Treatment of protracted diarrhea of infancy. 643 65

Amine-curing agent for epoxy resin, bis(4-amino-3-methylcyclohexyl)methane, has been suspected of inducing toxic symptoms in man which resemble collagen disease such as scleroderma or polymyositis. We studied subacute toxicity of this agent by repeated oral administrations to rats. The agent was administered 25, 37.5, 50, 75 or 100 mg/kg per one dose, 8 times in 10 days or 17 times in 4 weeks or 20-22 times in 4 weeks. When administration had been completed, clinical biochemical tests and histopathological examinations were carried out. Animals of high dosage group showed suppression of body weight increase and loss of muscular strength of limbs in the administration period. By clinical biochemical tests, elevation of blood components from muscle (CPK, GOT, creatine) was noticed. Also, increase of monoamine oxidase and decrease of alkaline phosphatase were revealed. By histological examinations, skeletal muscle and choroid plexus of brain showed noticeable changes. In muscles of high dosage group, atrophy, degeneration and regeneration of muscle fibers were observed and an increase of fibroblasts was also seen. In choroid plexus, vacuolar changes in epithelial cells were observed, being clearly dose-dependent. No particular change was recognized in skin. Though scleroderma-like change was not observed in the skin histologically, our results suggest that this amine-curing agent was greatly associated with muscle symptoms in the workers who handled epoxy resin and that it was one of causative agents which induced toxic symptoms like those of collagen disease.
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PMID:[Subacute toxicity of an amine-curing agent for epoxy resin]. 652 Oct 56

The present work describes an assay which is highly specific for ribose-5-phosphate. The method is based on the following three-stage enzymatic conversion: (1) ribose 5-phosphate in equilibrium ribose 1-phosphate (phosphopentomutase); (2) ribose 1-phosphate + adenine in equilibrium adenosine + Pi (adenosine phosphorylase); (3) adenosine + H2O----inosine + NH3 (adenosine deaminase). Ribose 5-phosphate may be determined either directly following the change in absorbance at 265 nm associated with the conversion of adenine to inosine, or radioenzymatically by measuring the radioactivity of inosine formed from [8-14C]adenine, after chromatographic separation of the nucleoside on polyethyleneimine-cellulose. The spectrophotometric assay was used to follow ribose 5-phosphate formation and ribose 1-phosphate consumption catalyzed by phosphopentomutase. Further, the ability of alkaline phosphatase, 5'-nucleotidase and crude extract of Bacillus cereus cells to act on ribose 5-phosphate was tested. The radioenzymatic assay was proved useful in determining the levels of ribose 5-phosphate in rat tissues.
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PMID:Spectrophotometric and radioenzymatic determination of ribose-5-phosphate. 653 May 7

The effect of cimetidine on abnormally elevated serum levels of parathyroid hormone was studied in 21 patients with secondary hyperparathyroidism due to chronic renal failure, receiving regular dialysis treatment. The concentrations of carboxyl (-COOH) and aminoterminal (-NH2) fragments of circulating immunoreactive parathyroid hormone (iPTH) were determined before and after 2 months af treatment with cimetidine 400 mg/day. All patients had, on admission, raised levels of either hormonal fragment. The mean pre-treatment value was 17.2 mU/ml for -COOH terminus (upper normal limit 6.5 mU/ml) and 5.7 mU/ml for -NH2 terminus (upper normal limit 1.9 mU/ml). At the end of cimetidine treatment the mean values were 19.9 and 5.7 mU/ml respectively for the two forms of circulating iPTH. No changes in total serum calcium, phosphate or alkaline phosphatase activity were recorded during the study. These results do not indicate any lowering effect of cimetidine on serum iPTH in chronic uraemic patients with secondary hyperparathyroidism.
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PMID:Effect of cimetidine on serum carboxyl and aminoterminal fragments of parathyroid hormone in chronic uraemic patients. 669 46

A comparative clinical trial of Travasol 8.5% without electrolytes and FreAmine II 8.5% was performed. Thirty-six patients were admitted to the double-blind study and assigned randomly to receive either Travasol (16 patients) or FreAmine II (20 patients). All patients had abnormal renal and liver function. Nitrogen balance was compared, and metabolic complications were monitored by evaluating BUN, serum creatinine, creatinine clearance, serum CO2, SGOT, SGPT, serum LDH, and serum alkaline phosphatase. There were no significant differences found in nitrogen balance of patients receiving either Travasol or FreAmine (p greater than 0.05). The Bun for days 1--5 in the Travasol group was significantly greater (p = 0.02), but the difference (3 mg/dl) was not considered clinically significant. All other comparative measurements of renal function, liver function, and acid-base balance remained the same throughout the study period (p greater than 0.05). Based on these results, the use of either FreAmine II or Travasol can promote positive nitrogen balance in a TPN patient population with normal renal and hepatic function with no difference in metabolic complications.
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PMID:Clinical comparison of two 8.5% amino acid injection products. 678 69

The complete amino acid sequence of the Escherichia coli alkaline phosphatase subunit [orthophosphoric-monoester phosphohydrolase (alkaline optimum), EC 3.1.3.1, isozyme 3] has been determined. The monomer contains 449 amino acid residues in a single unglycosylated polypeptide chain having a calculated Mr of 47,029. Isozyme 1 has an additional arginine residue at the NH2 terminus that presumably results from variability in processing of precursor molecules. Sequence data were obtained from both manual and automatic Edman degradation of the tryptic and cyanogen bromide peptides, as well as other peptides derived therefrom. The two disulfide bonds were determined from analyses of the appropriate peptic peptides. This structure confirms earlier reports of the sequence surrounding the active-site serine and both the NH2- and COOH-terminal cyanogen bromide fragments. A secondary structure prediction places nearly half the residues in alpha-helical segments that have 13% and 16%, respectively, in beta-strand and beta-turn orientations.
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PMID:Amino acid sequence of Escherichia coli alkaline phosphatase. 702 51

1. A study has been carried out on the steady-state kinetics followed by the alkaline phosphatase from Escherichia coli at different pH, temperatures, ionic strengths, phosphate concentrations and in the presence of the effectors such as Tris, NH4+--NH3 and CH3OH; p-nitrophenyl phosphate was used as substrate. 2. Contrary to what has generally been accepted, in most cases the enzyme follows non-Michaelian kinetics for a wide substrate concentration range, giving concave-down Lineweaver-Burk plots. Only at high phosphate concentrations (5 . 10(-3) M) and at high ionic strengths (2.0 M) is a linear Lineweaver-Burk plot obtained (Michaelian kinetics). 3. In order to analyse the kind of kinetics obtained, a non-linear regression fitting method was used to obtain rate vs substrate concentration equations as polynomial quotients of minimum degree with positive coefficients. 4. Most of the data obtained follows 2:2 degree type equations. 5. These results tend to suggest an idea of cooperativity rather than one of independence between the sites of the dimeric enzyme. A model is discussed for cooperativity between the sites with a wide concentration range giving concave-down Lineweaver-Burk plots.
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PMID:Negative cooperativity in alkaline phosphatase from E. col: new kinetic evidence from a steady-state study. 704 Jan 34

Three young dogs with a history of apathy, anorexia and weight loss were presented with severe ascites. Abnormal laboratory findings include hypoalbuminaemia and increased activities of alkaline phosphatase, serum aspartate amino-transferase, serum glutamic pyruvic transaminase and gamma-glutamyl transferase. Ammonia tolerance was also abnormal. At autopsy ascites and peripheral portosystemic collaterals were found. The livers were abnormally small and firm and their surfaces were irregular. Histologically, there was marked periportal fibrosis, increased numbers of bile ductules and arteriolae in the portal areas and an absence of normal portal vein tributaries. No inflammatory changes were found. These lesions are discussed in relation to the various causes of hepatic fibrosis.
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PMID:Hepatoportal fibrosis in three young dogs. 711 76


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