scholarly journals The expression of catechol oxidase activity during the hydroxylation of p-coumaric acid by spinach-beet phenolase

1972 ◽  
Vol 127 (4) ◽  
pp. 641-647 ◽  
Author(s):  
P. F. T. Vaughan ◽  
V. S. Butt
Keyword(s):  
Oxygen Consumption ◽  
Bovine Serum Albumin ◽  
Oxygen Uptake ◽  
Caffeic Acid ◽  
Bovine Serum ◽  
Acid Production ◽  
Reaction Rates ◽  
Catechol Oxidase ◽  
Coumaric Acid ◽  
Enzyme Preparations

1. The conditions under which oxygen consumption in excess of that required for the hydroxylation of p-coumaric acid to caffeic acid, catalysed by spinach-beet phenolase, can be suppressed, have been examined. 2. With dimethyltetrahydropteridine as electron donor, oxygen uptake was exactly equivalent to the caffeic acid produced, provided that p-coumaric acid was in excess, but with excess of reductant, oxygen uptake caused by the further oxidation of caffeic acid was also observed. 3. With equal concentrations of ascorbate and p-coumaric acid, equivalent oxygen uptake and caffeic acid production was found only in the first stages of the reaction, whereas with NADH substituted for ascorbate, oxygen uptake was in excess throughout. 4. When ascorbate was used, the period of the reaction over which this equivalence was found was decreased at high reaction rates and not observed at all with aged enzyme preparations; equivalence was restored by adding bovine serum albumin to these aged preparations. 5. Equivalence between oxygen consumption and caffeic acid production was observed with NADH, if small quantities of dimethyltetrahydropteridine were also added. 6. It is concluded that hydroxylation proceeds without the concomitant production of caffeic acid only if the enzyme is stabilized for hydroxylation by p-coumaric acid and the reductant, and is protected from attack by o-quinones.

scholarly journals The action of o-dihydric phenols in the hydroxylation of p-coumaric acid by a phenolase from leaves of spinach beet (Beta vulgaris L.)

1970 ◽  
Vol 119 (1) ◽  
pp. 89-94 ◽  
Author(s):  
P. F. T. Vaughan ◽  
V. S. Butt
Keyword(s):  
Caffeic Acid ◽  
Beta Vulgaris ◽  
Oxidase Activity ◽  
Maximum Rate ◽  
Specific Activity ◽  
Catechol Oxidase ◽  
Coumaric Acid ◽  
Catalytically Active ◽  
Lag Period ◽  
High Concentrations

1. Under defined conditions, the hydroxylation of p-coumaric acid catalysed by a phenolase from leaves of spinach beet (Beta vulgaris L.) was observed to develop its maximum rate only after a lag period. 2. By decreasing the reaction rate with lower enzyme concentrations or by increasing it with higher concentrations of reductants, the length of the lag period was inversely related to the maximum rate subsequently developed. 3. Low concentrations of caffeic acid or other o-dihydric phenols abolished this lag period. With caffeic acid, the rate of hydroxylation was independent of the reductant employed. 4. Hydroxylation was inhibited by diethyldithiocarbamate, but with low inhibitor concentrations hydroxylation recovered after a lag period. This lag could again be abolished by the addition of high concentrations of caffeic acid or other o-dihydric phenols. 5. Catechol oxidase activity showed no lag period, and did not recover from diethyldithiocarbamate inhibition. 6. The purified enzyme contained 0.17–0.33% copper; preparations with the highest specific activity were found to have the highest copper content. 7. The results are interpreted to suggest that the oxidation of o-dihydric phenols converts the enzymic copper into a species catalytically active in hydroxylation. This may represent the primary function for the catechol oxidase activity of the phenolase complex. The electron donors are concerned mainly, but not entirely, in the reduction of o-quinones produced in this reaction.

scholarly journals The hydroxylation of p-coumaric acid by an enzyme from leaves of spinach beet (Beta vulgaris L.)

1969 ◽  
Vol 113 (1) ◽  
pp. 109-115 ◽  
Author(s):  
P. F. T. Vaughan ◽  
V S Butt
Keyword(s):  
Sodium Chloride ◽  
Caffeic Acid ◽  
Beta Vulgaris ◽  
Catechol Oxidase ◽  
Purification Procedure ◽  
Coumaric Acid ◽  
Constant Ratio ◽  
Molar Basis ◽  
Tetrahydrofolic Acid ◽  
High Concentrations

1. An enzyme from the leaves of spinach beet (Beta vulgaris L.) that catalyses the hydroxylation of p-coumaric acid to caffeic acid in the presence of ascorbate has been purified about 1000-fold on a protein basis. 2. It is activated by high concentrations of ammonium sulphate and sodium chloride. 3. The preparation shows both hydroxylase and catechol oxidase activities, in a constant ratio throughout the purification procedure; they are similarly activated by salts. 4. Ascorbate acts as a reductant in quantities equivalent to the caffeic acid produced by hydroxylation. 5. Ascorbate can be replaced by tetrahydrofolic acid, NADH, NADPH or 2-amino-4-hydroxy-6,7-dimethyl-5,6,7,8-tetrahydropteridine, but not by caffeic acid. Among these, the pteridine is the most effective, but the reaction is not inhibited by aminopterin. In experiments with saturating concentrations of NADH and the pteridine, these reductants compete in the reaction and are equivalent on a molar basis. 6. No cofactor has been separated from the enzyme by prolonged dialysis. 7. The relation of the enzyme to other hydroxylases and phenolases is discussed.

In vitro study of caffeic acid — bovine serum albumin interaction

1988 ◽  
Vol 13 (1) ◽  
pp. 11-14 ◽  
Author(s):  
T. Adzet ◽  
J. Camarasa ◽  
E. Escubedo ◽  
M. Merlos
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Caffeic Acid ◽  
Bovine Serum ◽  
In Vitro Study ◽  
Vitro Study

Immunocytochemical localization of phenolic compounds in pollen walls using antibodies againstp-coumaric acid coupled to bovine serum albumin

PROTOPLASMA ◽  
1997 ◽  
Vol 197 (3-4) ◽  
pp. 148-159 ◽  
Author(s):  
C. Niester-Nyveld ◽  
A. Haubrich ◽  
H. Kampendonk ◽  
S. Gubatz ◽  
K. B. Tenberge ◽  
...  
Keyword(s):  
Phenolic Compounds ◽  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Bovine Serum ◽  
Immunocytochemical Localization ◽  
Coumaric Acid

scholarly journals Properties of rat liver microsomal stearoyl-coenzyme A desaturase

1977 ◽  
Vol 161 (2) ◽  
pp. 431-437 ◽  
Author(s):  
R Jeffcoat ◽  
P R Brawn ◽  
R Safford ◽  
A T James
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Rat Liver ◽  
Bovine Serum ◽  
Desaturase Activity ◽  
Cytochrome B5 ◽  
Fatty Acyl ◽  
Enzyme Preparations ◽  
Stearoyl Coa Desaturase ◽  
Nadh Cytochrome

1. Rat liver microsomal stearoyl-CoA desaturase activity was shown to be stimulated by both bovine serum albumin and a basic cytoplasmic protein from rat liver. 2. Partially purified desaturase is unaffected by either of these two proteins. 3. Bovine serum albumin appears to exert its effect on the crude system by protecting the desaturase substrate, stearoly-CoA, from the action of endogenous thiolesterases. 4. By using partially purified enzyme preparations, it was possible to establish the substate specificity of the delta9-fatty acyl-CoA desaturase with the C14, C15, C16, C17, C18 and C19 fatty acyl-CoA substrates. Maximum enzyme activity was shown with stearoyl-CoA decreasing with both palmitoyl-CoA and nonadecanoyl-CoA, as reported previously for free fatty acids. 5. Both cytochrome b5 and NADH-cytochrome b5 reductase (EC 1.6.2.2) are required for these studies and a method is described for the purification of homogeneous preparations of detergent-isolated cytochrome b5 from rat liver. 6. From amino acid analyses, a comparison was made of the hydrophobicity of the membrane portion of cytochrome b5 with the hydrophobicity reported for stearoyl-CoA desaturase. The close resemblance of the two values suggested that unlike cytochrome b5 and its reductase, the stearoyl-CoA desaturase may be largely buried in the endoplasmic reticulum.

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