Superoxide-forming NADPH oxidase preparation of pig polymorphonuclear leucocytes

A phagocytic vesicle fraction with high NADPH-dependent superoxide-forming activity was obtained in large quantity from pig blood polymorphonuclear leucocytes, phagocytosing oil droplets in the presence of cyanide. Essentially all of the superoxide-forming activity was extracted by repeated extracti...

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Published inBiochemical journal Vol. 205; no. 3; pp. 593 - 601
Main Authors Wakeyama, H, Takeshige, K, Takayanagi, R, Minakami, S
Format Journal Article
LanguageEnglish
Published 01.01.1982
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Abstract A phagocytic vesicle fraction with high NADPH-dependent superoxide-forming activity was obtained in large quantity from pig blood polymorphonuclear leucocytes, phagocytosing oil droplets in the presence of cyanide. Essentially all of the superoxide-forming activity was extracted by repeated extraction with a mixture containing deoxycholate and Tween 20. The extract had a superoxide-forming activity of 1 mu mol/min per mg of protein with NADPH, and one-fifth of this with NADH, K sub(m) values being similar to those of the vesicle fraction (40 mu M for NADPH and 400 mu M for NADH). A stoichiometric relationship of 1:2 for NADPH oxidation and superoxide formation was obtained, in agreement with the reaction NADPH + 2O sub(2) arrow right NADP super(+) + 2O sub(2) super(-) super(.) + H super(+). The activity of the extract was enhanced 2-fold by the addition of FAD, suggesting that the flavin is a component of the enzyme system. The K sub(m) value for FAD was 0 multiplied by 077 mu M.
AbstractList A phagocytic vesicle fraction with high NADPH-dependent superoxide-forming activity was obtained in large quantity from pig blood polymorphonuclear leucocytes, phagocytosing oil droplets in the presence of cyanide. Essentially all of the superoxide-forming activity was extracted by repeated extraction with a mixture containing deoxycholate and Tween 20. The extract had a superoxide-forming activity of 1 mu mol/min per mg of protein with NADPH, and one-fifth of this with NADH, K sub(m) values being similar to those of the vesicle fraction (40 mu M for NADPH and 400 mu M for NADH). A stoichiometric relationship of 1:2 for NADPH oxidation and superoxide formation was obtained, in agreement with the reaction NADPH + 2O sub(2) arrow right NADP super(+) + 2O sub(2) super(-) super(.) + H super(+). The activity of the extract was enhanced 2-fold by the addition of FAD, suggesting that the flavin is a component of the enzyme system. The K sub(m) value for FAD was 0 multiplied by 077 mu M.
Author Minakami, S
Takayanagi, R
Wakeyama, H
Takeshige, K
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Snippet A phagocytic vesicle fraction with high NADPH-dependent superoxide-forming activity was obtained in large quantity from pig blood polymorphonuclear leucocytes,...
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SubjectTerms leukocytes (polymorphonuclear)
NADPH oxidase
pigs
purification
Title Superoxide-forming NADPH oxidase preparation of pig polymorphonuclear leucocytes
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