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Location of the Epitope for 7D5, a Monoclonal Antibody Raised against Human Flavocytochrome b 558 , to the Extracellular Peptide Portion of Primate gp91 phox
Author(s) -
Yamauchi Akira,
Yu Lixin,
Pötgens Andy J.G.,
Kuribayashi Futoshi,
Nunoi Hiroyuki,
Kanegasaki Shiro,
Roos Dirk,
Malech Harry L.,
Dinauer Mary C.,
Nakamura Michio
Publication year - 2001
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.2001.tb02614.x
Subject(s) - epitope , biology , monoclonal antibody , p22phox , microbiology and biotechnology , linear epitope , epitope mapping , antibody , extracellular , protein subunit , biochemistry , gene , genetics
Flavocytochrome b 558 is the membrane component of the phagocyte NADPH oxidase, and is a heterodimer composed of gp91 phox and p22 phox subunits. Human flavocytochrome b 558 is recognized by monoclonal antibody 7D5 at an unidentified extracellular domain, although our previous study suggested it might recognize p22 phox . 7D5 has proven useful in rapid screening of individuals for X‐linked chronic granulomatous disease by flow‐cytometry. Therefore, we re‐evaluated the location of the 7D5 epitope using gene‐engineered cell lines expressing hybrid flavocytochromes composed of human and murine subunit homologues. The current study demonstrates that the 7D5 recognizes epitope only of primate gp91 phox . Flow‐cytometric analyses showed that 7D5 consistently bound to cells expressing human gp91 phox . In addition, 7D5 immunoprecipitated the ~58 kDa unglycosylated gp91 phox protein from solubilized membrane fractions of tunicamycin‐treated PLB‐985 granulocytes, indicating that glycans were not required for 7D5 binding. Transgenic COS7 cells expressing human gp91 phox but not p22 phox were recognized by 7D5. These results localized the epitope of 7D5 to an extracellular peptide portion of primate gp91 phox and indicate that the antibody will be useful for monitoring the efficiency of gene therapy in patients with flavocytochrome b 558 ‐deficient chronic granulomatous disease and for elucidating structural characteristics of flavocytochrome b 558 .