Bovine heart cardiolipin

Horse heart cyt c (type VI, oxidized form), bovine heart cardiolipin (>80% polyunsatured fatty acid content, primary linoleic acid; 98% purity), and laurdan were provided from Sigma Aldrich- Merck KGaA (Darmstadt, Germany) and used without further purification. All reagents were analytical grade.
For mutant cyt c the expression plasmids of horse cyt c (pHCyc) [50 (link)] was subjected to one round of mutagenesis with QuikChange Site-Directed Mutagenesis Kit (Agilent Technologies, Santa Clara United States), which introduced Lys72Asn substitution into the horse cyt c gene. Mutant pHCyc plasmids was introduced into E. coli JM 109. Protein expression and purification of the recombinant protein were then conducted as previously described [51 (link)].

1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), 1-oleoyl-2-hydroxy-sn-glycero-3-phosphocholine (OPC), bovine heart cardiolipin (CL), arachidonic acid (AA), hexane, hexadecane, KCl, HEPES, EDTA, Na₂SO₄, MES, Tris, adenosine 5-triphosphate (ATP), and guanosine 5-triphosphate (GTP) were purchased from Sigma-Aldrich (Munich, Germany). MPC (1-myristoyl-2-hydroxy-sn-glycero-3-phosphocholine) was obtained from Avanti Polar Lipids (Alabaster, AL, USA). Plain silica beads (22-μm diameter) were from Microspheres-Nanospheres (Corpuscular Co., Cold Spring, NY, USA). Chloroform was purchased from Carl Roth (Karlsruhe, Germany). 4-oxo-trans-2-nonenal (4-ONE) and 4-hydroxy-trans-2-nonenal (4-HNE) were purchased from Cayman Chemical (Ann Arbor, MI, USA) or synthesized as previously described [45 (link)].