Cloning and sequencing of rabbit leukocyte NADPH oxidase genes reveals a unique p67phoxhomolog

Abstract

The NADPH oxidase plays an important role in immune and nonimmune cell functions. Because rabbits represent an established model for studying a number of important disease processes that involve NADPH oxidase activity, we carried out studies to clone and sequence all five rabbit leukocyte NADPH oxidase genes. Comparison of the rabbit sequences with those of other species showed that, with the exception of p67phox, the rabbit phox proteins were highly conserved. In contrast, rabbit p67phox had a very divergent C-terminus and was 17 amino acids longer than any other known p67phox homolog. This was surprising, given the high degree of conservation among all of the phox proteins sequenced previously. To evaluate the functional consequences of this difference, wild-type rabbit p67phox and a mutated rabbit p67phox missing the C-terminal 17 amino acids were expressed and analyzed in a cell-free assay. Our results show that the full-length and truncated rabbit p67phox proteins were able to support oxidase activity, although the truncated form reproducibly supported a higher level of activity than full-length p67phox. These studies contribute to our understanding of the nature of the leukocyte NADPH oxidase in different species and will be valuable in future research using the rabbit model.