scholarly journals Partial purification and some properties of a cholinesterase from bush bean (Phaseolus vulgaris L.) roots

1978 ◽  
Vol 175 (3) ◽  
pp. 769-777 ◽  
Author(s):  
D H Mansfield ◽  
G Webb ◽  
D G Clark ◽  
I E P Taylor
Keyword(s):  
Phaseolus Vulgaris ◽  
Specific Activity ◽  
Apparent Molecular Weight ◽  
Fold Increase ◽  
Partial Purification ◽  
Phaseolus Vulgaris L ◽  
Native Form ◽  
Bush Bean ◽  
Electric Eel ◽  
Stokes Radius

A cholinesterase was partially purified from bush bean (Phaseolus vulgaris L.) roots by using acridinium-based ligand affinity chromatography. The procedure gave a 78-fold increase in specific activity, although at least three inactive contaminants remained. The enzyme activity was maximal against acetyl esters of choline and was inhibited by neostigmine. Di-isopropyl phosphorofluoridate completely inhibited activity at concentrations greater than 0.1 mM. The catalytic centre activity was 2 × 10(-4) times that of electric eel acetylcholinesterase. Cholinesterase activity appeared as a peak (s = 4.2 +/- 0.1 S) after isokinetic sedimentation. The Stokes radius was 4.00 nm and the apparent molecular weight was 72700 +/- 1900. The smallest active and native form of the enzyme appeared to be a monomer. This contrasts with animal acetylcholinesterases, in which the smallest active and native forms are multimeric.

TISSUE DISTRIBUTION AND BIOCHEMICAL CHARACTERIZATION OF BETA-CYANOALANINE SYNTHASE FROM COMMON BEAN SEEDS (PHASEOLUS VULGARIS)

2018 ◽  
Vol 6 ◽  
pp. 92
Author(s):  
Esther Nkechi Ezima ◽  
Adedeji Nelson Ademakinwa ◽  
Temitope Abigail Fasanya ◽  
Ayodeji Adebayo Adelegan ◽  
Abodunrin Emmanuel Ojo
Keyword(s):  
Phaseolus Vulgaris ◽  
Tissue Distribution ◽  
Biochemical Characterization ◽  
Developmental Stages ◽  
Specific Activity ◽  
Gel Filtration ◽  
Apparent Molecular Weight ◽  
Partial Purification ◽  
Chloride Salt ◽  
Cyanoalanine Synthase

Cyanide is produced throughout a plant's life cycle, its production increases during certain developmental stages such as seed germination, seedling elongation, fruit ripening and senescence. Beta-cyanoalanine synthase is the most important cyanide metabolizing enzyme found in plants. This work is aimed at studying the tissue distribution, partial purification and some physicochemical properties of β-cyanoalanine synthase from bean seeds (Phaseolus vulgaris). β-cyanolalanine synthase was isolated and partially purified using a combination of ammonium sulphate precipitation, desalting on Sephadex G-10 and gel filtration chromatography on Sephacryl S-200 column. The biochemical characteristics of the enzyme were investigated. Results obtained from this work showed that β-cyanolalanine synthase is more concentrated in the seeds (20.53 nmol/HS/mg) when compared 2 to the cotyledons (10.08 nmol/HS/mg) and the seed coats (5.82 nmol/HS/mg). The partially purified 2 2 enzyme showed a specific activity of 26.77 nmol/HS/mg and an apparent molecular weight of about 2 60,000Da, K values for cyanide and L-cysteine of  0.741 mM and 1.724 mM respectively. The V max m value obtained for cyanide was 25.00 nmol/H S/ml/minwhile that of L-cysteine was 2 o666.67nmol/H S/ml/min. The enzyme showed an optimum temperature of 40C and optimum pH of 2 10.0.Studies on the effect of chloride salt indicated that NaCl and MnCl had strong inhibitory effect 2 on the enzyme; NHCl had slight negative effect while KCl and ZnCl activated the enzyme dose 4 2 dependently.This study showed the presence of β-cyanoalanine synthase in bean seeds which is believed to function in the detoxification of cyanide produced in its tissues especially during germination.

Changes in the peroxidase activity of subcellular fractions from aging Phaseolus hypocotyls

1975 ◽  
Vol 53 (9) ◽  
pp. 852-860 ◽  
Author(s):  
B. Truelove ◽  
R. Rodriguez-Kabana ◽  
Larry R. Jones
Keyword(s):  
Phaseolus Vulgaris ◽  
Peroxidase Activity ◽  
Specific Activity ◽  
Total Activity ◽  
Chronological Age ◽  
Phaseolus Vulgaris L ◽  
Soluble Peroxidase ◽  
Tissue Homogenates ◽  
Hypocotyl Tissue ◽  
Nitrogen Contents

Changes in nitrogen contents and peroxidase activities of fractions isolated from hypocotyl tissue of Black Valentine bean (Phaseolus vulgaris L.) of increasing age were studied. As beans aged in darkness, a decreasing percentage of their nitrogen content was recovered in the isolated particulate fractions. Peroxidase activity of particulate fractions from dark-grown beans accounted for 49% of the total activity of both 3-day-old seedlings and 16-day-old senescing plants. Peroxidase specific activity of dark-grown tissue homogenates did not increase with plant age; however, after a certain period of growth, further aging resulted in increased peroxidase specific activity associated with the particulate fractions. Between day 3 and day 8 the patterns of peroxidase activity of the different fractions varied, but over the period day 9 to day 16, the patterns of all fractions were correlated. The nitrogen contents and peroxidase activities of fractions isolated from beans transferred from dark to light were different from those of fractions from beans of similar chronological age kept in darkness. Transfer of plants to light resulted in increased soluble peroxidase activity and prevention of the steep increase in particulate fraction activity recorded for dark-grown plants.

scholarly journals Hepatic microsomal Ca2+-dependent ATPase. Calmodulin-dependence and partial purification

1983 ◽  
Vol 214 (1) ◽  
pp. 69-75 ◽  
Author(s):  
P B Moore ◽  
N Kraus-Friedmann
Keyword(s):  
Affinity Chromatography ◽  
Microsomal Fraction ◽  
Specific Activity ◽  
Fold Increase ◽  
Partial Purification ◽  
Dependent Atpase

The hepatic microsomal fraction contains tightly bound calmodulin as demonstrated by affinity chromatography. When this calmodulin was partially removed by EGTA treatment (0.5 mM-EGTA), the uptake of 45Ca2+ by the microsomal vesicles was stimulated by added calmodulin and inhibited by trifluoperazine (TFP). The Ca2+-dependent ATPase was partially purified on a calmodulin column. This partial purification resulted in a 500-fold increase in the specific activity of the enzyme when measured in the presence of added calmodulin. Antibodies prepared against calmodulin prevented this stimulatory effect. The fraction eluted from the calmodulin column contained several protein bands indicating that the specific activity of the Ca2+-dependent ATPase is probably still underestimated. There are likely to be other calmodulin-sensitive processes present in the hepatic microsomal fraction.

Partial purification of a hypertensive substance from rat erythrocytes

1985 ◽  
Vol 63 (10) ◽  
pp. 1321-1326 ◽  
Author(s):  
William D. McCumbee ◽  
Gary L. Wright
Keyword(s):  
Blood Pressure ◽  
Systolic Blood Pressure ◽  
Calcium Uptake ◽  
Specific Activity ◽  
Present Report ◽  
Fold Increase ◽  
Partial Purification ◽  
Rat Erythrocytes ◽  
In Vitro Effects

A crude extract prepared from rat erythrocytes was previously shown to contain an active component (hypertensive factor, HF) that stimulates the in vitro uptake of calcium by aortic rings and causes a sustained and severe elevation in systolic blood pressure in normotensive rats. The present report demonstrates that the in vitro effects of HF on calcium uptake are concentration dependent and reversible, and that these effects on calcium uptake provide a convenient bioassay for monitoring the purification of HF. HF was prepared from a diffusate obtained by dialyzing hemolyzed rat erythrocytes. The diffusate was applied successively to molecular sieve and anion exchange columns resulting in a 1500-fold increase in specific activity relative to the starting hemolysate. The compound stimulating calcium uptake was found to be heat stable and resistant to digestion with trypsin and chymotrypsin. In contrast, treatment with pronase E, a nonspecific protease, markedly increased the calcium transport stimulatory activity. The administration of the purified preparation to normotensive rats having a systolic blood pressure of 118 ± 2 Torr (1 Torr = 133.322 Pa) resulted in a significant elevation of blood pressure (171 ± 11 Torr) by day 5. The severity of this increase further suggests that there is a marked enhancement in potency relative to earlier fractions that were examined. Perhaps most importantly, these results indicate that, at this stage of purification, the pressor component and the calcium-stimulatory component of HF appear to co-purify.

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