Structural basis for cytochrome c Y67H mutant to function as a peroxidase.

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Cytochrome c (cyt c) catalyzes cardiolipin peroxidation, essential for apoptosis.
• The structural basis for cyt c's peroxidase activity is not fully understood.

Purpose of the Study:

• To determine the 3D NMR solution structure of a yeast cyt c Y67H variant with high peroxidase activity.
• To elucidate the structural and mechanistic basis of cardiolipin peroxidation by cyt c.

Main Methods:

• Three-dimensional NMR spectroscopy to determine the structure of yeast cyt c Y67H variant.
• Steady-state kinetic studies to assess peroxidase activity at different pH conditions.

Main Results:

• The Y67H variant structure is similar to native cyt c, with a disrupted hydrogen bond between Met80 and residue 67.
• This disruption destabilizes the heme Fe(3+)-Met80 bond, making it labile at low pH.
• Optimal peroxidase activity for the Y67H variant occurs between pH 4.0 and 5.2.

Conclusions:

• The His67 residue in the Y67H variant functions as a distal histidine.
• Protonation of His67 facilitates H2O2 cleavage, acting as an acidic catalyst in cardiolipin peroxidation.
• A mechanism for cardiolipin peroxidation catalyzed by the Y67H variant is proposed.