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Immunological studies on the respiratory burst oxidase of pig blood neutrophils
Author(s) -
Fukuhara Yukiko,
Ise Yayoi,
Kakinuma Katsuko
Publication year - 1988
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(88)80816-0
Subject(s) - isoelectric focusing , microbiology and biotechnology , enzyme , polyacrylamide gel electrophoresis , biochemistry , respiratory burst , chemistry , guinea pig , antibody , oxidase test , gel electrophoresis , flavin group , enzyme assay , biology , immunology , endocrinology
Recently, a flavin enzyme (p I 5.0), that is probably responsible for superoxide (O − 2 )‐generating oxidase activity, was separated by isoelectric focusing‐polyacrylamide gel electrophoresis (IEF—PAGE) from neutrophil membranes in our laboratory [(1987) J. Biol. Chem. 262, 12316–12322]. In the present work, we performed immunological studies on this enzyme derived from pig blood neutrophils. The enzyme extract obtained on IEF—PAGE was injected into guinea pigs to raise antibodies. IgG antibody against the p I 5.0 protein inhibited maximally 54% of the O − 2 ‐generating activity of the membrane‐solubilized oxidase, whereas the normal serum IgG was not inhibitory at all. Our results further confirmed that the enzyme (p I 5.0) is one of the component(s) of the O − 2 ‐generating system. The enzyme gave rise to a band corresponding to a major protein of 72±4 kDa on both non‐denaturing and SDS—PAGE. Immunoblotting after SDS—PAGE demonstrated labelling of peptides of 70–72, 28–32 and 16–18 kDa.