Cytochrome aa3 from Nitrosomonas europaea

Area of Science:

• Biochemistry
• Microbiology
• Enzyme Kinetics

Background:

• Nitrosomonas europaea is an ammonia-oxidizing bacterium crucial for nitrification.
• Cytochrome c oxidase plays a vital role in cellular respiration and energy production.
• Previous classification of this enzyme as 'cytochrome a1' was based on limited spectral data.

Purpose of the Study:

• To purify and characterize the cytochrome c oxidase from Nitrosomonas europaea.
• To determine the enzyme's structure, metal content, and spectral properties.
• To re-evaluate its classification based on new experimental evidence.

Main Methods:

• Purification using ion-exchange chromatography with Triton X-100.
• Spectroscopic analysis (absorption maxima in resting and reduced states).
• EPR spectroscopy to analyze metal centers and enzyme states.
• Protein quantification using the Lowry procedure.

Main Results:

• The purified enzyme exhibits absorption maxima characteristic of cytochrome aa3.
• It contains approximately 11 nmol heme a and 11 nmol copper per mg protein.
• EPR spectra are consistent with a mitochondrial cytochrome aa3-type oxidase.
• The enzyme is composed of three subunits with distinct molecular weights.

Conclusions:

• The enzyme from Nitrosomonas europaea is definitively classified as cytochrome aa3, not cytochrome a1.
• This reclassification is supported by its metal content and EPR spectral properties.
• The natural electron donor for this enzyme remains to be identified, though horse heart cytochrome c is a rapid reductant.