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Cytochrome aa3 from Nitrosomonas europaea.

1986; Elsevier BV; Volume: 261; Issue: 36 Linguagem: Inglês

10.1016/s0021-9258(19)75997-3

1083-351X

Alan A. DiSpirito, John D. Lipscomb, Alan B. Hooper,

Mass Spectrometry Techniques and Applications

Cytochrome c oxidase has been purified from the ammonia oxidizing chemoautotroph Nitrosomonas eu- ropaea by ion-exchange chromatography in the presence of Triton X-100.The enzyme has absorption maxima at 420 and 592 nm in the resting state and at 444 and 598 nm in the dithionite-reduced form; optical extinction coefficient (598 nm minus 640 nm) = 21.9 cm" m"'.The enzyme has -1 1 nmol of heme a and -11 nmol of copper per mg of protein (Lowry procedure).There appear to be three subunits (approximate molecular weights 50,800, 38,400, and 35,500), two heme groups (a and a3), and two copper atoms per minimal unit.The EPR spectra of the resting and partially reduced enzyme are remarkably similar to the corresponding spectra of the mitochondrial cytochrome aadype oxidase.Although the enzyme had been previously classified as "cytochrome a," OQ the basis of its ferrous a absorption maximum (598 nm), its metal content and EPR spectral properties clearly show that it is better classified as a cytochrome as.Neither the data reported here nor a review of the literature supports the existence of cytochrome al as an entity discrete from cytochrome a a 3 .The purified enzyme is reduced rapidly by ferrous horse heart cytochrome c or cytochrome c-554 from N. europaea, but not with cytochrome e-552 from N .europaea.The identity of the natural electron donor is as yet unestablished.With horse heart cytochrome c as electron donor, the purified enzyme could account for a significant portion of the terminal oxidase activity in vivo.Heme a-containing terminal oxidases from bacteria have traditionally been classified as either cytochrome a a 3 or cytochrome al according to their absorption spectrum.Reduced cytochromes with absorbance maxima at 440-445 and 600-605 nm in the Soret and (Y regions, respectively, have been designated cytochrome a a 3 , while maxima at 435-445 and 585-595 nm are taken to be characteristic of cytochrome a1 (1-4).Several enzymes of each type have been reported.Representatives of one group of cytochrome aa3-type oxidases that have been purified from bacteria exhibit enzymatic and spectral properties similar to those of mitochondrial cyto-