Defining the Apoptotic Trigger

O’Brien, Evan S.; Nucci, Nathaniel V.; Fuglestad, Brian; Tommos, Cecilia; Wand, A. Joshua

doi:10.1074/jbc.m115.689406

Cited by 53 publications

(40 citation statements)

References 54 publications

(74 reference statements)

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“…In particular, the rupture of the heme-Fe-M80 coordination and the tertiary rearrangement of cyt c, involving the 40's Ω loop and the M80-containing Ω loop, have been reported to occur in CL-bound cyt c [7,20,21]. The difficulties encountered in the study of the cyt c/CL binding process are correlated with several factors, such as the ionic strength of the solution, the membrane curvature, the CL/ cyt c molar ratio at which the binding reaction is followed, which significantly affect the results obtained [7,17,19,45,46]. Moreover, the finding that CL-bound cyt c is present in solution as a heterogeneous ensemble of forms characterized by different heme coordination represents a further complication [15,16,28].…”

Section: Discussionmentioning

confidence: 99%

The key role played by charge in the interaction of cytochrome c with cardiolipin

et al. 2016

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mutations cancel the CL-dependent peroxidase activity of cyt c; furthermore, CL does not interact with the Lys72Asn mutant. In the present paper, we extend our study to the Lys → Arg mutants to investigate the influence exerted by the charge possessed by the residues located at positions 72 and 73 on the cyt c/CL interaction. On the basis of the present work a number of overall conclusions can be drawn: (i) position 72 must be occupied by a positively charged residue to assure cyt c/CL recognition; (ii) the Arg residues located at positions 72 and 73 permit cyt c to react with CL; (iii) the replacement of Lys72 with Arg weakens the second (low-affinity) binding transition; (iv) the Lys73Arg mutation strongly increases the peroxidase activity of the CL-bound protein.Abstract Cytochrome c undergoes structural variations upon binding of cardiolipin, one of the phospholipids constituting the mitochondrial membrane. Although several mechanisms governing cytochrome c/cardiolipin (cyt c/CL) recognition have been proposed, the interpretation of the process remains, at least in part, unknown. To better define the steps characterizing the cyt c-CL interaction, the role of Lys72 and Lys73, two residues thought to be important in the protein/lipid binding interaction, were recently investigated by mutagenesis. The substitution of the two (positively charged) Lys residues with Asn revealed that such Electronic supplementary material The online version of this article

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Section: Discussionmentioning

confidence: 99%

The key role played by charge in the interaction of cytochrome c with cardiolipin

et al. 2016

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“…1 In addition to its key role as an electron carrier in the electron-transport chain, cyt c is also instrumental in apoptosis by mediating oxidation of cardiolipin (CL) in mitochondrial membranes 2 – 4 and, upon its release to cytosol, binding to Apaf-1 to form an apoptosome. An edge of the heme in cyt c is partially exposed to the solvent and multiple Lys residues surround the heme crevice.…”

Section: Introductionmentioning

confidence: 99%

The K79G Mutation Reshapes the Heme Crevice and Alters Redox Properties of Cytochromec

et al. 2018

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The two roles of cytochrome c (cyt c), in oxidative phosphorylation and apoptosis, critically depend on redox properties of its heme iron center. The K79G mutant has served as a parent protein for a series of mutants of yeast iso-1 cyt c. The mutation preserves the Met80 coordination to the heme iron, as found in WT* (K72A/C102S), and many spectroscopic properties of K79G and WT* are indistinguishable. The K79G mutation does not alter the global stability, fold, rate of Met80 dissociation, or thermodynamics of the alkaline transition (pKa) of the protein. However, the reduction potential of the heme iron decreases; further, the pKH of the trigger group and the rate of the Met-to-Lys ligand exchange associated with the alkaline transition decrease, suggesting changes in the environment of the heme. The rates of electron self-exchange and bimolecular electron transfer (ET) with positively-charged inorganic complexes increase, as does the intrinsic peroxidase activity. Analysis of the reaction rates suggests that there is increased accessibility of the heme edge in K79G and supports the importance of the Lys79 site for bimolecular ET reactions of cyt c, including those with some of its native redox partners. Structural modeling rationalizes the observed effects to arise from changes in the volume of the heme pocket and solvent accessibility of the heme group. Kinetic and structural analyses of WT* characterize the properties of the heme crevice of this commonly employed reference variant. This study highlights the important role of Lys79 for defining functional redox properties of cyt c.

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“…Site N (novel site), identified via chemical shift perturbation ( 15 N- 1 H-HSQC) upon titration of CL into Cyt c encapsulated in reverse micelles, encompasses residues Phe36, Gly37, Thr58, Trp59 and Lys60, and thus also likely involves a component of electrostatic binding. 14 Site C (cardiolipin site) is a hydrophobic binding site and thus is most relevant to the current study. Binding data for site C suggest that one or more of the fatty acyl chains of CL rotates out of the membrane and inserts into a hydrophobic channel in Cyt c .…”

Section: Introductionmentioning

confidence: 99%

Cytochrome c Can Form a Well-Defined Binding Pocket for Hydrocarbons

et al. 2016

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Cytochrome c can acquire peroxidase when it binds to cardiolipin in mitochondrial membranes. The resulting oxygenation of cardiolipin by cytochrome c provides an early signal for the onset of apoptosis. The structure of this enzyme-substrate complex is a matter of considerable debate. We present three structures at 1.7 – 2.0 Å resolution of a domain-swapped dimer of yeast iso-1-cytochrome c with the detergents, CYMAL-5, CYMAL-6 and ω-undecylenyl-β-D-maltopyranoside bound in a channel that places the hydrocarbon moieties of these detergents next to the heme. The heme is poised for peroxidase activity with water bound in place of Met80, which serves as the axial heme ligand when cytochrome c functions as an electron carrier. The hydroxyl group of Tyr67 sits 3.6 – 4.0 Å from the nearest carbon of the detergents, positioned to act as a relay in radical abstraction during peroxidase activity. Docking studies with linoleic acid, the most common fatty acid component of cardiolipin, show that C11 of linoleic acid can sit adjacent to Tyr67 and the heme, consistent with the oxygenation pattern observed in lipidomics studies. The well-defined hydrocarbon binding pocket provides atomic resolution evidence for the extended lipid anchorage model for cytochrome c/cardiolipin binding. Dimer dissociation/association kinetics for yeast versus equine cytochrome c indicate that formation of mammalian cytochrome c dimers in vivo would require catalysis. However, the dimer structure shows that only a modest deformation of monomeric cytochrome c would suffice to form the hydrocarbon binding site occupied by these detergents.

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Defining the Apoptotic Trigger

Cited by 53 publications

References 54 publications

The key role played by charge in the interaction of cytochrome c with cardiolipin

The key role played by charge in the interaction of cytochrome c with cardiolipin

The K79G Mutation Reshapes the Heme Crevice and Alters Redox Properties of Cytochromec

Cytochrome c Can Form a Well-Defined Binding Pocket for Hydrocarbons

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