J Vet Sci.  2000 Jun;1(1):27-31.

Activation domain in P67phox regulates the steady state reduction of FAD in gp91phox

• Han CH
• Lee MH

Abstract

An activation domain in p67(phox) (residues 199-210) is critical for regulating NADPH oxidase activity in cell-free system [10] To determine the steady state reduction of FAD, thioacetamide-FAD was reconstituted in gp91(phox), and the fluorescence of its oxidised form was monitored. Omission of p67(phox) decreased the steady state reduction of the FAD from 28% to 4%, but omission of p47(phox) had little effect. A series of the truncated forms of p67(phox) were expressed in E.coli to determine the domain in p67(phox) which is essential for regulating the steady state of FAD reduction. The minimal length of p67(phox) for for regulating the steady state of FAD reduction is shown to be 1-210 using a series of truncation mutants which indicates that the region 199-210 is also important for regulating electron flow within flavocytochrome b(558). The deletion of this domain not only decreased the superoxide generation but also decreased the steady state of FAD reduction. Therefore, the activation domain on p67(phox) regulates the reductive half-reaction for FAD, consistent with a dominant effect on hydride/electron transfer from NADPH to FAD.

Keyword

the activation domain on p67(phox); the steady state of FAD reduction

MeSH Terms

Amino Acid Sequence
Base Sequence
Cell Membrane/metabolism
Cell-Free System
DNA Primers
Flavin-Adenine Dinucleotide/*metabolism
Humans
Kinetics
Membrane Glycoproteins/*metabolism
Molecular Sequence Data
NADH Dehydrogenase/metabolism
*NADPH Oxidase
Neutrophils/enzymology/metabolism
Oxidation-Reduction
Peptide Fragments/chemistry
Phosphoproteins/*chemistry/*metabolism
Polymerase Chain Reaction
Sequence Deletion