Nitric Oxide Binding to the Cardiolipin Complex of Ferric Cytochrome c

Abstract

Cardiolipin, a phospholipid specific to the mitochondrion, interacts with the small electron transfer heme protein cytochrome c through both electrostatic and hydrophobic interactions. Once in a complex with cardiolipin, cytochrome c has been shown to undergo a conformational change that leads to the rupture of the bond between the heme iron and the intrinsic sulfur ligand of a methionine residue and to enhance the peroxidatic properties of the protein considered important to its apoptotic activity. Here we report that the ferric cytochrome c/cardiolipin complex binds nitric oxide tightly through a multistep process in which the first step is the relatively slow displacement (5 s(-1)) from heme coordination of an intrinsic ligand that replaces methionine in the complex. Nanosecond photolysis of the nitrosyl adduct demonstrated that a fraction of the nitric oxide escapes from the heme pocket and subsequently recombines to the heme in second-order processes (k = 1.8 × 10(6) and 5.5 × 10(5) M(-1) s(-1)) that, under these conditions, were much faster than recombination of the intrinsic ligand with which they compete. Ultrafast (femtosecond) laser photolysis showed that the geminate recombination of nitric oxide to the heme occurred with time constants (τ = 22 and 72 ps) and that ~23% of the photolyzed nitric oxide escaped into the bulk phase. This high value for the escape fraction relative to other heme proteins indicates the open nature of the heme pocket in this complex. These results are summarized in a scheme and are discussed in terms of the possible modulation of the apoptotic activity of cytochrome c by nitric oxide.

NO2−-mediated nitrosylation of ferrous microperoxidase-11

“…Secondly, MP11-Fe represents a heme-model compound for highly reactive penta-coordinated CM-cyt c and CL-cyt c [7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22], although it is not a physiological heme-protein. Remarkably, CL-cyt c, displaying myoglobin-like spectroscopic and ligand binding properties [7][8][9][10][11][12][13][14][15][16][17][18][19][20][21], is pivotal for switching cyt c functions from mitochondrial respiration to apoptosis.…”

“…Remarkably, CL-cyt c, displaying myoglobin-like spectroscopic and ligand binding properties [7][8][9][10][11][12][13][14][15][16][17][18][19][20][21], is pivotal for switching cyt c functions from mitochondrial respiration to apoptosis. The partial unfolding of cyt c upon interaction with CL is accompanied by changes in its chemical reactivity including the enzymatic activity.…”

“…The fifth coordination ligand of the heme-Fe atom of cyt c is invariantly His, whereas the sixth coordination ligand exhibits high variability, being Met in horse heart cyt c [1][2][3][4][5][6][7]. The heme-Fe atom of hexa-coordinated native horse heart cyt c is essentially unreactive (or very low reactive), whereas the heme-Fe atom of penta-coordinated cyt c derivatives (i.e., carboxymethylated cyt c (CM-cyt c), cardiolipinbound CM-cyt c (CL-CM-cyt c), and cardiolipin-bound cyt c (CL-cyt c)) displays myoglobin-like spectroscopic and functional properties [7][8][9][10][11][12][13][14][15][16][17][18][19][20][21].…”

“…1), the sixth heme-Fe coordination ligand being the O atom of a water molecule in the ferric form. Therefore, MPs are heme-model compounds of choice to highlight the modulatory role of the protein matrix and of the heme-Fe atom axial ligation on the spectroscopic, ligand binding, and (pseudo-)enzymatic properties of horse heart cyt c. In particular, MPs represent heme-model compounds of highly reactive penta-coordinated CM-cyt c, CL-CM-cyt c, and CL-cyt c derivatives, the last being involved in apoptosis initiation [7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22].…”

Structural changes and picosecond to second dynamics of cytochrome c in interaction with nitric oxide in ferrous and ferric redox states

“…74 The ferric Cyt c-NO association rate thus appears of the same order of magnitude as other ferric heme proteins 75 and especially is the same as in ferric nitrophorin but ten times faster than in ferric Mb. In the presence of cardiolipin bound to the ferric Cyt c two bimolecular rebinding components were observed, 42 ). This suggests that one of these species in the presence of cardiolipin keeps the native conformation of ferric Cyt c. ).…”