Activation studies by phospholipids on the purified cytochrome c4:o oxidase of Azotobacter vinelandii.

A modified procedure is described that was used to solubilize and purify the TMPD-dependent cytochrome c4:o oxidase from Azotobacter vinelandii. Two functional components (Fractions I and V) were obtained after DEAE-cellulose chromatography. Fraction V contained both cytochrome c4 (3.6 nmol/mg protein) and cytochrome o (1.6 nmol/mg protein). This cytochrome oxidase complex oxidized TMPD at "moderate" rates. Fraction I, a clear greenish-yellow fraction, contained primarily phosphatidylethanolamine with some phosphatidylglycerol. Fraction I itself could not oxidize TMPD, but when it was preincubated with Fraction V, a 2-4-fold stimulation in TMPD oxidase activity occurred. Other "authentic" micellar phospholipids also readily activated TMPD oxidase activity in Fraction V. The maximum activation effect obtained with Fraction I was in essence duplicated with purified phosphatidylethanolamine.