scholarly journals Dihydro-orotase from Clostridium oroticum. Purification and reversible removal of essential zinc

1985 ◽  
Vol 230 (1) ◽  
pp. 101-108 ◽  
Author(s):  
D W Pettigrew ◽  
R R Bidigare ◽  
B J Mehta ◽  
M I Williams ◽  
E G Sander
Keyword(s):  
Enzyme Activity ◽  
Amino Acid ◽  
Phosphate Buffer ◽  
Specific Activity ◽  
Zinc Content ◽  
Atomic Absorption Spectrophotometry ◽  
Kinetic Properties ◽  
Purification Procedure ◽  
Normal Complement ◽  
Absorption Spectrophotometry

A new purification procedure involving five column-chromatography steps is described for dihydro-orotase (L-5,6-dihydro-orotate amidohydrolase, EC 3.5.2.3) from Clostridium oroticum (A.T.C.C. 25750). The native purified enzyme is a dimer of Mr 102 000 and contains 4.0 +/- 0.3 g-atoms of zinc/mol of dimer. These observations agree with those reported previously [Taylor, Taylor, Balch & Gilchrist (1976) J. Bacteriol. 127, 863-873]. It is conclusively demonstrated that dihydro-orotase is a zinc metalloenzyme. Zinc is reversibly removed by treatment with chelators in phosphate buffer at pH 6.5, as demonstrated by atomic absorption spectrophotometry and decrease of enzyme activity. The specific activity is linearly dependent on zinc content. Addition of ZnSO4 to the chelator-treated enzyme results in regain of the normal complement of zinc and enzyme activity. Kinetic properties of the reconstituted enzyme are indistinguishable from those of the native enzyme. The amino acid composition of the homogeneous enzyme suggests that the zinc atoms occupy different environments.

Direct Measurement of the Hepatointestinal Extraction of Zinc in Cirrhosis and Hepatitis

1981 ◽  
Vol 61 (4) ◽  
pp. 441-444 ◽  
Author(s):  
P. W. N. Keeling ◽  
W. Ruse ◽  
J. Bull ◽  
B. Hannigan ◽  
R. P. H. Thompson
Keyword(s):  
Chronic Hepatitis ◽  
Statistical Significance ◽  
Venous Blood ◽  
Zinc Content ◽  
Atomic Absorption Spectrophotometry ◽  
Zinc Extraction ◽  
Blood Samples ◽  
Peripheral Venous Blood ◽  
Absorption Spectrophotometry ◽  
Biopsy Specimens

1. 65Zn was injected intravenously during transjugular liver biopsy and, from simultaneous hepatic and peripheral venous blood samples, hepatointestinal 65Zn extraction was calculated. Hepatic zinc content was measured in biopsy specimens. 2. On the same occasion samples of liver tissue were taken and their zinc content was measured by atomic absorption spectrophotometry. 3. Seven patients with cirrhosis had significantly lower hepatic zinc content and hepatointestinal zinc extraction than six control patients with mild liver disease. Six patients with chronic hepatitis had a mean hepatointestinal zinc extraction higher than control patients, whereas their mean hepatic zinc content was lower, although the former difference did not achieve statistical significance. 4. These results demonstrate that hepatointestinal extraction of zinc is impaired in cirrhosis, but not in chronic hepatitis.

scholarly journals Escherichia coli S-adenosylhomocysteine/5′-methylthioadenosine nucleosidase. Purification, substrate specificity and mechanism of action

1985 ◽  
Vol 232 (2) ◽  
pp. 335-341 ◽  
Author(s):  
F Della Ragione ◽  
M Porcelli ◽  
M Cartenì-Farina ◽  
V Zappia ◽  
A E Pegg
Keyword(s):  
Escherichia Coli ◽  
Enzyme Activity ◽  
Mechanism Of Action ◽  
Specific Activity ◽  
Maximal Rate ◽  
Purification Procedure ◽  
Protein Affinity ◽  
Naturally Occurring ◽  
Purine Ring ◽  
Ribose Moiety

S-Adenosylhomocysteine/5′-methylthioadenosine nucleosidase (EC 3.2.2.9) was purified to homogeneity from Escherichia coli to a final specific activity of 373 mumol of 5′-methylthioadenosine cleaved/min per mg of protein. Affinity chromatography on S-formycinylhomocysteine-Sepharose is the key step of the purification procedure. The enzyme, responsible for the cleavage of the glycosidic bond of both S-adenosylhomocysteine and 5′-methylthioadenosine, was partially characterized. The apparent Km for 5′-methylthioadenosine is 0.4 microM, and that for S-adenosylhomocysteine is 4.3 microM. The maximal rate of cleavage of S-adenosylhomocysteine is approx. 40% of that of 5′-methylthioadenosine. Some 25 analogues of the two naturally occurring thioethers were studied as potential substrates or inhibitors of the enzyme. Except for the analogues modified in the 5′-position of the ribose moiety or the 2-position of the purine ring, none of the compounds tested was effective as a substrate. Moreover, 5′-methylthioformycin, 5′-chloroformycin, S-formycinylhomocysteine, 5′-methylthiotubercidin and S-tubercidinylhomocysteine were powerful inhibitors of the enzyme activity. The results obtained allow the hypothesis of a mechanism of enzymic catalysis requiring as a key step the protonation of N-7 of the purine ring.

scholarly journals Diversity of Iron and Zinc Content in Bananas from East and Central Africa

HortScience ◽  
2016 ◽  
Vol 51 (4) ◽  
pp. 320-324
Author(s):  
Michael Pillay ◽  
Robert Fungo
Keyword(s):  
Central Africa ◽  
Germplasm Collection ◽  
Zinc Content ◽  
Atomic Absorption Spectrophotometry ◽  
Absorption Spectrophotometry ◽  
Fiber Protein ◽  
Fe Content ◽  
Zn Content ◽  
Wide Variability ◽  
Iron And Zinc

Bananas and plantains (Musa sp.) are major staple foods in many developing countries of the world. Although bananas are rich in carbohydrate, fiber, protein, fat, and vitamins A, C, and B6 they are largely deficient of iron (Fe), iodine, and zinc (Zn). A small increase in the micronutrient content of bananas could play a major role in combating disorders that are due to deficiency of mineral micronutrients such as Fe and Zn. The objective of this study was to determine the Fe and Zn content of 47 banana genotypes from a germplasm collection in Uganda using atomic absorption spectrophotometry. The Fe and Zn content showed wide variability and highly significant differences (P < 0.001) within and among the different banana categories selected for this study. The highest average Fe content (1.42 mg/100 g) was found in ‘Saba’ (ABB) while the least Fe content (0.06 mg/100 g) was found in ‘Kikundi’ (AAA). The highest average Zn content (1.21 mg/100 g) among the analyzed accessions was found in ‘Kivuvu’ (ABB) while Zn was not detectable in both ‘Kabucuragye’ (AAA) and ‘Grand Naine’ (AAA). Considering these figures, there is a greater than 20-fold variation in the Fe and Zn levels of the banana genotypes used this study suggesting that genetic improvement of genotypes for enhanced micronutrient levels may be achieved by breeding.

scholarly journals SYNTHESIS AND ANALYSIS OF ZINC METHIONINE, ZINC GLYCINE, COPPER LEUCINE, AND COPPER GLYCINE COMPLEXES USING ATOMIC ABSORPTION SPECTROPHOTOMETRY

2018 ◽  
Vol 10 (1) ◽  
pp. 388
Author(s):  
Sekar Alinda Nastiti ◽  
Harmita . ◽  
Catur Jatmika
Keyword(s):  
Amino Acid ◽  
Atomic Absorption ◽  
Acid Synthesis ◽  
Atomic Absorption Spectrophotometry ◽  
Water Soluble ◽  
Synthesis Process ◽  
Mineral Concentration ◽  
Amino Acid Synthesis ◽  
Absorption Spectrophotometry ◽  
Free Metal

Objective: The aim of this study was to perform metal-amino acid synthesis and to analyze the free and bonded mineral concentrations.Methods: In this study, the synthesis of amino acid metal complexes was carried out by reacting free metal ions, derived from a water-soluble metalsalt, with amino acids in a 1:2 molar ratio.Results: The respective yields of this synthesis process were 95.38%, 95.95%, 76.31%, and 93.91% for zinc (Zn)-methionine (Zn(Met)2), Zn-glycine(Zn(gli)2), copper-leucine (Cu(leu)2), and Cu-glycine (Cu(gli)2) complexes, respectively. The metal-amino acid complexes were then separated usingcolumn chromatography and further analyzed by atomic absorption spectrophotometry (AAS). The bonded metal concentrations of the Zn(Met)2,Zn(gli)2, Cu(leu)2, and Cu(gli)2 complexes were 189.32 mg/g, 353.78 mg/g, 180.89 mg/g, and 275.11 mg/g, respectively. The free metal concentrationsof the Zn(Met)2, Zn(gli)2, Cu(leu)2, and Cu(gli)2 complexes were 13.57 mg/g, 12.92 mg/g, 0.19 mg/g, and 2.12 mg/g, respectively.Conclusion: In this study, Zn(Met)2, Zn(gli)2, Cu(leu)2, and Cu(gli)2 complexes were successfully formed and analyzed. The mineral concentration ineach complex differed depending on the type of mineral and ligand.

Influence of Ions and Chelating Agents on the Haemolymphacetylcholinesterase of Mytilus edulis

1978 ◽  
Vol 33 (11-12) ◽  
pp. 851-858 ◽  
Author(s):  
Dettmar von Wachtendonk ◽  
Jutta Neef
Keyword(s):  
Enzyme Activity ◽  
Mytilus Edulis ◽  
Chelating Agents ◽  
Competitive Inhibition ◽  
Divalent Cations ◽  
Atomic Absorption Spectrophotometry ◽  
Ammonium Ions ◽  
Salicylic Aldehyde ◽  
Inorganic Cations ◽  
Absorption Spectrophotometry

By use of different inhibitors as well as atomic absorption spectrophotometry it has been shown that the haemolymph-acetylcholinesterase (E. c. 3.1.1.7) of the sea mussel Mytilus edulis is a me- talloprotein containing 2,95 Fe2+ -ions per subunit. All inhibitors used (1,10-phenanthroline, salicylic aldehyde, 2,2′-dipyridyl, 8-hydroxyquinoline) showed a non-competitive inhibition, which was not pH- dependent. Some divalent cations caused a marked increase of the enzyme activity, some heavy metals inhibited the enzyme almost completely; monovalent inorganic cations did not influence the enzyme at all. Besides NaF and Na2SiF6, which showed a non-competitive inhibition comparable to the inhibition observed with the chelating agents, and NaN3 , whose mode of action was not identifiable, no inhibition by different mono- and divalent inorganic anions was to be observed. Ammonium ions caused no enzyme inhibition, but length the inhibition power of substituted ammonium ions increased with an increasing C-chain. The influence of some organic solvents on the enzyme activity is demonstrated.

ZINC AND MANGANESE CONTENTS OF MICRO-DISSECTED PANCREATIC ISLETS OF SOME RODENTS

1977 ◽  
Vol 86 (3) ◽  
pp. 570-577 ◽  
Author(s):  
Niilo Havu ◽  
Gillis Lundgren ◽  
Sture Falkmer
Keyword(s):  
Heavy Metals ◽  
Guinea Pig ◽  
Pancreatic Islets ◽  
Guinea Pigs ◽  
Graphite Furnace ◽  
Chinese Hamster ◽  
Zinc Content ◽  
Wistar Rat ◽  
Atomic Absorption Spectrophotometry ◽  
Absorption Spectrophotometry

ABSTRACT By a flameless atomic absorption spectrophotometry procedure, using a graphite furnace, it was possible to assay the contents of zinc and manganese in micro-dissected pancreatic islets of several rodents. Interest was focused upon the islets of guinea-pigs, due to the fact that guinea-pig insulin lacks a histidine residue in the B10 position of the molecule which normally binds zinc (or other heavy metals) in the hexamer formation, probably involved in the storage of insulin. Both the zinc and manganese contents were too low in the guinea-pig islet parenchyma to be reasonably involved in the storage of insulin in the β-granules. Instead, it was suggested that guinea-pig insulin, like hagfish insulin, might crystallize without access to zinc or other heavy metals. Low zinc and manganese contents were also observed in newborn and diabetic guinea-pigs. The islet zinc content was high in the Wistar rat, the Chinese hamster, and the spiny mouse. No significant amounts of manganese were found in any of these kinds of islet parenchyma.

Levels of hippocampal calcium and zinc following kindling-induced epilepsy

1985 ◽  
Vol 63 (2) ◽  
pp. 159-161 ◽  
Author(s):  
I. Mody ◽  
J. J. Miller
Keyword(s):  
Dentate Gyrus ◽  
Atomic Absorption ◽  
Granule Cells ◽  
Zinc Content ◽  
Dry Weight ◽  
Atomic Absorption Spectrophotometry ◽  
Absorption Spectrophotometry

Hippocampal calcium and zinc content was determined using atomic absorption spectrophotometry in control and commissural-kindled rats. In animals exhibiting 5–10 consecutive motor seizures hippocampal calcium was slightly elevated (356.7 parts per million (ppm), dry weight) but not significantly different from controls (329.8 ppm), whereas the amount of zinc was significantly higher (101.6 ppm) than in nonstimulated animals (88.3 ppm). These results are indicative of certain pathophysiological changes in kindled hippocampi, most likely localized to the granule cells of the dentate gyrus where the bulk of hippocampal zinc is confined.

scholarly journals Dopamine β-hydroxylase of bovine adrenal medullae. A rapid purification procedure

1972 ◽  
Vol 126 (5) ◽  
pp. 1209-1217 ◽  
Author(s):  
A. Foldes ◽  
P. L. Jeffrey ◽  
B. N. Preston ◽  
L. Austin
Keyword(s):  
Amino Acid ◽  
Specific Activity ◽  
Purification Procedure ◽  
Chromaffin Granules ◽  
Endogenous Inhibitor ◽  
Membrane Bound ◽  
Rapid Purification ◽  
The Stability ◽  
And Storage

1. A rapid purification procedure for dopamine β-hydroxylase from bovine adrenal-medulla chromaffin granules is presented. The homogeneity of the purified enzyme was demonstrated by means of three independent criteria. The specific activity of the enzyme compares favourably with that obtained by more involved procedures. 2. The stability of the enzyme was investigated and storage in polypropylene tubes was found preferable to storage in glass. 3. The soluble and particulate forms of dopamine β-hydroxylase appear to be identical, since membrane-bound and membrane-enclosed forms of the enzyme exhibit similar properties as regards size, charge and amino acid composition. 4. Ca2+ was found to stimulate the release of dopamine β-hydroxylase from bovine chromaffin granules in vitro. 5. An endogenous inhibitor of the enzyme was found in the chromaffin granules. This inhibitor was not inactivated either by heating at 100°C or by pretreatment with p-chloromercuribenzoate or Cu2+ ions.

THE CHEMICAL MODIFICATION OF CHYMOTRYPSIN

1964 ◽  
Vol 42 (6) ◽  
pp. 695-714 ◽  
Author(s):  
G. H. Dixon ◽  
H. Schachter
Keyword(s):  
Enzyme Activity ◽  
Amino Acid ◽  
Chemical Modification ◽  
Active Site ◽  
Specific Binding ◽  
Bond Breaking ◽  
Kinetic Properties ◽  
Side Chains ◽  
Amino Acid Side Chains

Chemical modification of chymotrypsin has led to the identification of several amino acid side-chains which are probably constituents of the active site of the enzyme. A single seryl and a single histidyl residue appear to cooperate in catalyzing the bond-breaking process while one or more tryptophanyl residues may be involved in the specific binding of substrate. Neither of the two methionyl residues is essential for enzyme activity although changes in kinetic properties occur when they are modified by oxidation or alkylation.

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