title: ATP specifically drives refolding of non-native conformations of cytochrome c.
abstract: An increasing body of evidence ascribes to misfolded forms of cytochrome c (cyt c) a role in pathophysiological events such as apoptosis and disease. Here, we examine the conformational changes induced by lipid binding to horse heart cyt c at pH 7 and study the ability of ATP (and other nucleotides) to refold several forms of unfolded cyt c such as oleic acid-bound cyt c, nicked cyt c, and acid denatured cyt c. The CD and fluorescence spectra demonstrate that cyt c unfolded by oleic acid has an intact secondary structure, and a disrupted tertiary structure and heme environment. Furthermore, evidence from the Soret CD, electronic absorption, and resonance Raman spectra indicates the presence of an equilibrium of at least two low-spin species having distinct heme-iron(III) coordination. As a whole, the data indicate that binding of cyt c to oleic acid leads to a partially unfolded conformation of the protein, resembling that typical of the molten globule state. Interestingly, the native conformation is almost fully recovered in the presence of ATP or dATP, while other nucleotides, such as GTP, are ineffective. Molecular modeling of ATP binding to cyt c and mutagenesis experiments show the interactions of phosphate groups with Lys88 and Arg91, with adenosine ring interaction with Glu62 explaining the unfavorable binding of GTP. The finding that ATP and dATP are unique among the nucleotides in being able to turn non-native states of cyt c back to native conformation is discussed in the light of cyt c involvement in cell apoptosis.
authors.affiliation: Dipartimento di Medicina Sperimentale e Scienze Biochimiche, Università di Roma Tor Vergata, via Montpellier 1, 00133 Rome, Italy.
authors.forename: Federica
authors.initials: F
authors.lastname: Sinibaldi
authors.forename: Giampiero
authors.initials: G
authors.lastname: Mei
authors.forename: Fabio
authors.initials: F
authors.lastname: Polticelli
authors.forename: M Cristina
authors.initials: MC
authors.lastname: Piro
authors.forename: Barry D
authors.initials: BD
authors.lastname: Howes
authors.forename: Giulietta
authors.initials: G
authors.lastname: Smulevich
authors.forename: Roberto
authors.initials: R
authors.lastname: Santucci
authors.forename: Franca
authors.initials: F
authors.lastname: Ascoli
authors.forename: Laura
authors.initials: L
authors.lastname: Fiorucci
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D001665:Binding Sites; D045304:Cytochromes c / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D016296:Mutagenesis; D009711:Nucleotides / Q000494:pharmacology; D019301:Oleic Acid / Q000378:metabolism; D011487:Protein Conformation / Q000187:drug effects; D017510:Protein Folding; D013059:Spectrum Analysis, Raman
chemical_list: D009711:Nucleotides; D019301:Oleic Acid; D000255:Adenosine Triphosphate; D045304:Cytochromes c
references.citation: Cell. 1996 Jul 12;86(1):147-57
references.pmid: 8689682
references.citation: J Biol Chem. 2001 Jan 5;276(1):153-8
references.pmid: 11027687
references.citation: Proteins. 1991;11(4):281-96
references.pmid: 1758883
references.citation: J Mol Biol. 1990 Jul 20;214(2):585-95
references.pmid: 2166170
references.citation: J Biol Chem. 1995 Feb 17;270(7):3197-202
references.pmid: 7852404
references.citation: J Inorg Biochem. 1997 Nov 15;68(3):211-4
references.pmid: 9352653
references.citation: Adv Protein Chem. 1995;47:83-229
references.pmid: 8561052
references.citation: Biochemistry. 2000 Jul 18;39(28):8234-42
references.pmid: 10889031
references.citation: Biochemistry. 1989 Oct 17;28(21):8609-13
references.pmid: 2481497
references.citation: J Biol Chem. 2001 Jun 1;276(22):19356-62
references.pmid: 11279142
references.citation: Biochemistry. 1997 Aug 12;36(32):9867-77
references.pmid: 9245419
references.citation: Nature. 1991 Dec 5;354(6352):408-10
references.pmid: 1956406
references.citation: Cell Death Differ. 1999 Jul;6(7):683-8
references.pmid: 10453079
references.citation: Protein Sci. 2003 Dec;12(12):2805-14
references.pmid: 14627740
references.citation: Science. 1997 Feb 21;275(5303):1132-6
references.pmid: 9027315
references.citation: Biochemistry. 2000 Oct 17;39(41):12632-8
references.pmid: 11027143
references.citation: Biochem J. 1991 May 1;275 ( Pt 3):569-73
references.pmid: 2039436
references.citation: J Biol Chem. 2000 May 26;275(21):16127-33
references.pmid: 10821864
references.citation: Biochem Biophys Res Commun. 1974 Sep 9;60(1):371-5
references.pmid: 4371200
references.citation: Eur Biophys J. 2003 Nov;32(7):599-613
references.pmid: 12768249
references.citation: Eur Biophys J. 1990;18(3):193-201
references.pmid: 2162758
references.citation: J Biol Chem. 1974 Oct 25;249(20):6434-8
references.pmid: 4472692
references.citation: Biochemistry. 1998 May 5;37(18):6402-9
references.pmid: 9572857
references.citation: Cell. 1997 Nov 14;91(4):479-89
references.pmid: 9390557
references.citation: Biochem J. 1991 Nov 1;279 ( Pt 3):781-6
references.pmid: 1659388
references.citation: J Mol Biol. 2003 Aug 1;331(1):37-43
references.pmid: 12875834
references.citation: Eur J Biochem. 2001 Aug;268(16):4537-43
references.pmid: 11502215
references.citation: Biochemistry. 1972 Oct 24;11(22):4120-31
references.pmid: 4343790
references.citation: Arch Biochem Biophys. 2000 Jul 15;379(2):331-6
references.pmid: 10898952
references.citation: J Inorg Biochem. 2004 Jun;98(6):1067-77
references.pmid: 15149817
references.citation: Eur J Biochem. 1999 Apr;261(2):379-91
references.pmid: 10215847
references.citation: Nat Med. 1998 Oct;4(10):1157-65
references.pmid: 9771749
references.citation: Biopolymers. 2000;57(2):77-84
references.pmid: 10766958
references.citation: Biochemistry. 2003 May 27;42(20):6149-58
references.pmid: 12755617
references.citation: Curr Opin Struct Biol. 1997 Feb;7(1):41-52
references.pmid: 9032064
references.citation: J Biol Inorg Chem. 2003 May;8(5):527-539
references.pmid: 12764601
references.citation: Biochemistry. 2003 Jun 24;42(24):7604-10
references.pmid: 12809517
references.citation: J Biol Inorg Chem. 2004 Dec;9(8):997-1006
references.pmid: 15503233
references.citation: Biochemistry. 1995 Feb 28;34(8):2686-93
references.pmid: 7873551
references.citation: Biochemistry. 2001 Aug 14;40(32):9460-8
references.pmid: 11583145
references.citation: Biochim Biophys Acta. 1968 Jan 22;154(1):84-90
references.pmid: 5689051
references.citation: Protein Sci. 1995 Apr;4(4):716-28
references.pmid: 7613469
references.citation: J Biol Chem. 2002 Mar 15;277(11):8822-6
references.pmid: 11781329
references.citation: Biochemistry. 1999 Mar 23;38(12):3599-609
references.pmid: 10090746
references.citation: J Biol Chem. 1999 Mar 5;274(10):6388-96
references.pmid: 10037730
references.citation: Science. 1998 Aug 28;281(5381):1309-12
references.pmid: 9721092
references.citation: J Biol Chem. 1996 Aug 2;271(31):18379-86
references.pmid: 8702481
references.citation: J Mol Biol. 2004 Oct 29;343(4):1095-109
references.pmid: 15476824
references.citation: J Biochem. 1969 Feb;65(2):259-67
references.pmid: 5771953
references.citation: Proteins. 2002 Nov 15;49(3):385-97
references.pmid: 12360528
references.citation: Biochemistry. 1990 Apr 10;29(14):3480-8
references.pmid: 2162192
references.citation: Gen Physiol Biophys. 1995 Feb;14(1):19-37
references.pmid: 8529863
references.citation: Biochem Cell Biol. 2000;78(6):675-81
references.pmid: 11206578
references.citation: J Biol Chem. 1999 Mar 5;274(10):6080-4
references.pmid: 10037689