CLONING AND SEQUENCING OF THE BOVINE FLAVOCYTOCHROME-B SUBUNIT PROTEINS, GP91-PHOX AND P22-PHOX - COMPARISON WITH OTHER KNOWN FLAVOCYTOCHROME-B SEQUENCES

Neutrophils play an essential role in the cellular defense of the bovi ne mammary gland and compromised leukocyte function has been linked to the development of bovine mastitis. During mastitis, large numbers of leukocytes migrate into the mammary tissues where they become activat ed, resulting in the assembly of neutrophil membrane and cytosolic pro teins to form a superoxide anion-generating complex Known as the NADPH oxidase, The key membrane-associated component of the NADPH oxidase i s flavocytochrome b, which is a heterodimer of p22-phox and gp91-phox. Currently, only the human, porcine, murine, and rattus p22-phox and t he human, porcine, and murine gp91-phox gene sequences are known, Beca use of the important role neutrophils play in bovine host defense, we carried out studies to clone, sequence, and analyze expression of bovi ne flavocytochrome b, Using polymerase chain reaction cloning techniqu es and a bovine spleen cDNA library we have cloned both of the bovine flavocytochrome b subunits, p22-phox and gp91-phox. Comparison of the bovine sequences with those of other species also revealed important i nformation regarding key structural features of gp91-phox and p22-phox , including location of putative glycosylation sites. This study great ly contributes to our understanding of the potential functional sites of the flavocytochrome b subunits as well as providing information tha t can be used to study the role of neutrophils in bovine inflammatory diseases such as mastitis.