BIOCHEMICAL AND IMMUNOCHEMICAL PROPERTIES OF B-LYMPHOCYTE CYTOCHROME B(558)

Like neutrophils, Epstein-Barr virus (EBV)-immortalized B lymphocytes express all constituents of the NADPH oxidase complex necessary to gen erate superoxide anion O-2(-). The NADPH oxidase activity in EBV-B lym phocytes is only 5% of that measured in neutrophils upon PMA stimulati on. Cytochrome b(558) is the sole redox membrane component of NADPH ox idase; it is the protein core around which cytosolic factors assemble in order to mediate oxidase activity. In the present study, we have co mpared the structural and functional properties of cytochrome b(558) f rom EBV-B lymphocytes and neutrophils. Cytochrome b(558) from EBV-B ly mphocyte plasma membrane, like that from neutrophils, is characterized by a heterodimeric structure with a highly glycosylated beta subunit, known as gp91-phox. While the amount of cytochrome b(558) recovered a fter purification from EBV-B lymphocytes (similar to 0.24 nmol from 10 (10) cells) was low compared to that recovered from neutrophils (simil ar to 10 nmol), the biochemical properties of purified cytochrome b(55 8) from both EBV-B lymphocytes and neutrophils were quite similar with respect to their differential spectra, redox potential, and FAD bindi ng site. Once cytochrome b(558) was extracted from the EBV-B lymphocyt e membrane, it was able to mediate, in a reconstituted system of O-2(- ) production the same oxidase turnover as that found for cytochrome b( 558) extracted from neutrophils. A comparison between membrane bound a nd soluble cytochrome b(558) suggested that the weak oxidase activity measured in intact EBV-B cells might be the result not only of the sma ll amount of expressed cytochrome b(558), but also of a defect of the activation process in lymphocyte membrane. (C) 1998 Elsevier Science B .V.