The reactivity of cytochrome <i>c</i> with soft ligands

Schejter, Abel; Plotkin, Batya; Vig, Ida

doi:10.1016/0014-5793(91)80292-b

Cited by 46 publications

(54 citation statements)

References 13 publications

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“…determined polarographically at pH 7.9 (25 mM Tris acetate) (18). Computer graphics were obtained by the HYDRA (19) (26), resulting in cleavage of the methionine sulfur-heme iron bond and in disappearance of the 695-nm band that is associated with ligation of a low-spin ferric heme iron by bivalent sulfur (27,28 (Fig. 2).…”

Section: Methodsmentioning

confidence: 99%

Structural significance of an internal water molecule studied by site-directed mutagenesis of tyrosine-67 in rat cytochrome c.

Luntz

Schejter

Garber

et al. 1989

Proc. Natl. Acad. Sci. U.S.A.

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The tyrosine-67 to phenylalanine mutated rat cytochrome c is similar to the unmutated protein in its spectral, reduction potential, and enzymic electron-transfer properties. However, the loss of the 695-nm band, characteristic of the ferric form of the normal low-spin physiologically active configuration, occurs 1.2 pH units higher on the alkaline side and 0.7 pH unit lower on the acid side. Similarly, the heme ironmethionine-80 sulfur bond is more stable to temperature, with the midpoint ofthe transition being 300C higher, corresponding to an increase in AH of 5 kcal/mol (1 cal = 4.184 J), partially mitigated by an increase of 11 entropy units in AS. Urea has only slightly different effects on the two proteins. These phenomena are best explained by considering that the loss of one of the three hydrogen-bonding side chains, tyrosine-67, asparagine-52, and threonine-78, which hold an internal water molecule on the "left, lower front" side of the protein [Takano, Biol. 153,, is sufficient to prevent its inclusion in the mutant protein, leading to a more stable structure, and, as indicated by preliminary proton NMR two-dimensional phase-sensitive nuclear Overhauser effect spectroscopy analyses, a reorganization of this area. This hypothesis predicts that elimination ofthe hydrogenbonding ability of residue 52 or 78 would also result in cytochromes c having similar properties. It is not obvious why the space-filling structure involving the internalized water molecule that leads to a destabilization energy of about 3 kcal/mol should be subject to extreme evolutionary conservation, when a more stable and apparently fully functional structure is readily available.

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Section: Methodsmentioning

confidence: 99%

Structural significance of an internal water molecule studied by site-directed mutagenesis of tyrosine-67 in rat cytochrome c.

Luntz

Schejter

Garber

et al. 1989

Proc. Natl. Acad. Sci. U.S.A.

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“…It has been suggested that the covalent attachment is a device to prevent heme loss by dissociation (8). It has also been argued that the thioether bonds increase binding affinity of methionine to the ferrous iron and thus contribute to the relatively high reduction potentials of some c-type cytochromes (9). Recently, the effects of losing both the covalent bonds to heme have been described in studies where the cytochrome c 552 from a thermophile Hydrogenobacter thermophilus was converted into a b-type cytochrome as a result of mutation of the sequence CXXCH to AXXAH (10).…”

mentioning

confidence: 99%

Loss of Either of the Two Heme-binding Cysteines from a Class I c-Type Cytochrome Has a Surprisingly Small Effect on Physicochemical Properties

Tomlinson

Ferguson

2000

Journal of Biological Chemistry

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Almost without exception, c-type cytochromes have heme covalently attached via two thioether linkages to the cysteine residues of a CXXCH motif. The reasons for the covalent attachment are not understood. Reported here is cytoplasmic expression in Escherichia coli of AXXCH and CXXAH variants of cytochrome c 552 from Hydrogenobacter thermophilus; remarkably, the single thioether bond proteins have, apart from an altered visible absorption spectrum, almost identical properties, including thermal stability and reduction potential, to the wild type CXXCH protein. In combination with previous work showing that an AXXAH variant of cytochrome c 552 is much less stable than the CXXCH form, it can be concluded that covalent attachment of heme via either of thioether bonds is sufficient to confer considerable stability and that these bonds contribute little to the setting of the reduction potential. The absence of AXXCH or CXXAH heme-binding motifs from bacterial cytochromes c may relate to the coexistence of the assembly pathway with that for formation of disulfide bonds in the bacterial periplasm.

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“…Further, it has been argued that the position of the imidazole relative to the porphyrin plane maximizes the back-bonding properties of the iron and consequently increases the strength of the heme iron-methionine sulfur bond (6). The heme plane, with pyrrole rings indicated in roman numerals, is viewed from the top front, slightly tilted toward the left, so that the imidazole (im) of histidine-18 on the right side is closer to the viewer than the sulfur (S) of methionine-80, on the left side of the central heme iron atom (Fe).…”

mentioning

confidence: 99%

Changing the invariant proline-30 of rat and Drosophila melanogaster cytochromes c to alanine or valine destabilizes the heme crevice more than the overall conformation.

Koshy

Luntz

Schejter

et al. 1990

Proc. Natl. Acad. Sci. U.S.A.

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Drosophila melanogaster and rat cytochromes c in which proline-30 was converted to alanine or valine were expressed in a strain of baker's yeast, Saccharomyces cerevisiae, where they sustained aerobic growth. The mutations had no significant effect on the spectra or redox potentials but altered drastically the stability of the bond between the methionine-80 sulfur and the heme iron, as judged by four criteria: (i) the alkaline pKa values of the 695-nm band of the ferric form of the mutant proteins decreased by almost 1 pH unit as compared to the wild types; (ii) the acid pKa values increased by 0.5 to 1.2 pH units; (iii) the 695-nm band half-disappeared at temperatures 10-20TC lower in the mutant proteins than in the wild types; and (iv) the 695-nm band of the mutant proteins was susceptible to concentrations of urea that had little influence on their overall structure. The valinesubstituted rat cytochrome c had properties intermediate between those of the wild type and the alanine mutant. The destabilized coordinative bond is located in space a long distance from the mutation site. It is suggested that the mutations weaken the hydrogen bond between the carbonyl of residue 30 and the imino group of the imidazole of histidine-18, modifying the bonding of the heme iron by that imidazole, which, in turn, through a trans effect, weakens the bond between the heme iron and the other axial ligand, the sulfur of methionine-80. Alternatively, the effect of the mutations may be propagated allosterically along the peptide chain.

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The reactivity of cytochrome c with soft ligands

Cited by 46 publications

References 13 publications

Structural significance of an internal water molecule studied by site-directed mutagenesis of tyrosine-67 in rat cytochrome c.

Structural significance of an internal water molecule studied by site-directed mutagenesis of tyrosine-67 in rat cytochrome c.

Loss of Either of the Two Heme-binding Cysteines from a Class I c-Type Cytochrome Has a Surprisingly Small Effect on Physicochemical Properties

Changing the invariant proline-30 of rat and Drosophila melanogaster cytochromes c to alanine or valine destabilizes the heme crevice more than the overall conformation.

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