pH Dependence of Rate Constants for Reactions of Cytochrome c with Inorganic Redox Partners and Mechanistic Implications

P. L. Drake; R. T. Hartshorn; Joseph Mcginnis; A. G. Sykes

doi:10.1021/ic00306a029

pH Dependence of Rate Constants for Reactions of Cytochrome c with Inorganic Redox Partners and Mechanistic Implications

P. L. Drake, R. T. Hartshorn, Joseph Mcginnis, A. G. Sykes

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Abstract

Rate constants for the oxidation of horse cytochrome c(II) with [Co(phen)₃]³⁺and [Co(terpy)₂]³⁺decrease (~30%) on decreasing the pH from 8 to 5, giving a pK_avalue of 6.7 (average). No corresponding decrease is observed with tuna cytochrome c(II) (which has His26 but no His33) or with His33 diethyl pyrocarbonate (DEPC) modified horse cytochrome c(II). Candida krusei cytochrome c(II) gives a pK_aof 6.9, which is likewise assigned to His33. No dependence on pH is observed with the negatively charged oxidants [Fe(CN)₆]^3_and [Co(dipic)₂]^-or surprisingly with [Co(terpy)₂]²⁺, [Ru(NH₃)₅py]²⁺, and [Co(sep)]²⁺as reductants for horse cytochrome c(III). Previous work using CDNP-modified horse cytochrome c derivatives has indicated that the positively charged redox partners [Co(phen)₃]³⁺(oxidant) and [Co(sep)]²⁺(reductant) react preferentially at site II on the front face of the molecule to the right-hand side of the solvent-accessible heme edge (the latter vertical with axially coordinated Met to the left), whereas negatively charged [Fe(CN)₆]^3-reacts at site III to the left-hand side and across the top section of the exposed heme edge. His33 is close to site II, consistent with the effect of protonation of this residue on reactivity with the 3+ oxidants. Reduction potentials for horse cytochrome c determined electrochemically in the presence of a mediator show no variation with pH over the range 7.5-5. Protonation of His33, the imidazole ring of which is 12 A from Fe, has no effect on the reduction potential therefore, and its effect on reactivity is a local effect. It is not immediately clear why there is no dependence on pH for the reaction of cytochrome c(III) with the three positively charged reductants. Possible contributing factors include some sort of conformational change, with a resultant shift in the site for electron transfer in the case of for example [Co(terpy)₂]²⁺as compared to [Co(terpy)₂]³⁺, and effects stemming from different degrees of solvation at or near His33. His33 of horse cytochrome c gives virtually identical pA_a’s and rates of DEPC modification for both the oxidized and reduced forms, indicating similar degrees of accessibility.

Original language	English
Pages (from-to)	1361-1366
Number of pages	6
Journal	Inorganic Chemistry
Volume	28
Issue number	7
DOIs	https://doi.org/10.1021/ic00306a029
Publication status	Published - 1 Apr 1989

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@article{44a74061ce3e4eb58c0243dd56d1789e,

title = "pH Dependence of Rate Constants for Reactions of Cytochrome c with Inorganic Redox Partners and Mechanistic Implications",

abstract = "Rate constants for the oxidation of horse cytochrome c(II) with [Co(phen)3]3+and [Co(terpy)2]3+decrease (~30%) on decreasing the pH from 8 to 5, giving a pKavalue of 6.7 (average). No corresponding decrease is observed with tuna cytochrome c(II) (which has His26 but no His33) or with His33 diethyl pyrocarbonate (DEPC) modified horse cytochrome c(II). Candida krusei cytochrome c(II) gives a pKaof 6.9, which is likewise assigned to His33. No dependence on pH is observed with the negatively charged oxidants [Fe(CN)6]3_and [Co(dipic)2]-or surprisingly with [Co(terpy)2]2+, [Ru(NH3)5py]2+, and [Co(sep)]2+as reductants for horse cytochrome c(III). Previous work using CDNP-modified horse cytochrome c derivatives has indicated that the positively charged redox partners [Co(phen)3]3+(oxidant) and [Co(sep)]2+(reductant) react preferentially at site II on the front face of the molecule to the right-hand side of the solvent-accessible heme edge (the latter vertical with axially coordinated Met to the left), whereas negatively charged [Fe(CN)6]3-reacts at site III to the left-hand side and across the top section of the exposed heme edge. His33 is close to site II, consistent with the effect of protonation of this residue on reactivity with the 3+ oxidants. Reduction potentials for horse cytochrome c determined electrochemically in the presence of a mediator show no variation with pH over the range 7.5-5. Protonation of His33, the imidazole ring of which is 12 A from Fe, has no effect on the reduction potential therefore, and its effect on reactivity is a local effect. It is not immediately clear why there is no dependence on pH for the reaction of cytochrome c(III) with the three positively charged reductants. Possible contributing factors include some sort of conformational change, with a resultant shift in the site for electron transfer in the case of for example [Co(terpy)2]2+as compared to [Co(terpy)2]3+, and effects stemming from different degrees of solvation at or near His33. His33 of horse cytochrome c gives virtually identical pAa{\textquoteright}s and rates of DEPC modification for both the oxidized and reduced forms, indicating similar degrees of accessibility.",

author = "Drake, {P. L.} and Hartshorn, {R. T.} and Joseph Mcginnis and Sykes, {A. G.}",

year = "1989",

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doi = "10.1021/ic00306a029",

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journal = "Inorganic Chemistry",

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T1 - pH Dependence of Rate Constants for Reactions of Cytochrome c with Inorganic Redox Partners and Mechanistic Implications

AU - Drake, P. L.

AU - Hartshorn, R. T.

AU - Mcginnis, Joseph

AU - Sykes, A. G.

PY - 1989/4/1

Y1 - 1989/4/1

N2 - Rate constants for the oxidation of horse cytochrome c(II) with [Co(phen)3]3+and [Co(terpy)2]3+decrease (~30%) on decreasing the pH from 8 to 5, giving a pKavalue of 6.7 (average). No corresponding decrease is observed with tuna cytochrome c(II) (which has His26 but no His33) or with His33 diethyl pyrocarbonate (DEPC) modified horse cytochrome c(II). Candida krusei cytochrome c(II) gives a pKaof 6.9, which is likewise assigned to His33. No dependence on pH is observed with the negatively charged oxidants [Fe(CN)6]3_and [Co(dipic)2]-or surprisingly with [Co(terpy)2]2+, [Ru(NH3)5py]2+, and [Co(sep)]2+as reductants for horse cytochrome c(III). Previous work using CDNP-modified horse cytochrome c derivatives has indicated that the positively charged redox partners [Co(phen)3]3+(oxidant) and [Co(sep)]2+(reductant) react preferentially at site II on the front face of the molecule to the right-hand side of the solvent-accessible heme edge (the latter vertical with axially coordinated Met to the left), whereas negatively charged [Fe(CN)6]3-reacts at site III to the left-hand side and across the top section of the exposed heme edge. His33 is close to site II, consistent with the effect of protonation of this residue on reactivity with the 3+ oxidants. Reduction potentials for horse cytochrome c determined electrochemically in the presence of a mediator show no variation with pH over the range 7.5-5. Protonation of His33, the imidazole ring of which is 12 A from Fe, has no effect on the reduction potential therefore, and its effect on reactivity is a local effect. It is not immediately clear why there is no dependence on pH for the reaction of cytochrome c(III) with the three positively charged reductants. Possible contributing factors include some sort of conformational change, with a resultant shift in the site for electron transfer in the case of for example [Co(terpy)2]2+as compared to [Co(terpy)2]3+, and effects stemming from different degrees of solvation at or near His33. His33 of horse cytochrome c gives virtually identical pAa’s and rates of DEPC modification for both the oxidized and reduced forms, indicating similar degrees of accessibility.

AB - Rate constants for the oxidation of horse cytochrome c(II) with [Co(phen)3]3+and [Co(terpy)2]3+decrease (~30%) on decreasing the pH from 8 to 5, giving a pKavalue of 6.7 (average). No corresponding decrease is observed with tuna cytochrome c(II) (which has His26 but no His33) or with His33 diethyl pyrocarbonate (DEPC) modified horse cytochrome c(II). Candida krusei cytochrome c(II) gives a pKaof 6.9, which is likewise assigned to His33. No dependence on pH is observed with the negatively charged oxidants [Fe(CN)6]3_and [Co(dipic)2]-or surprisingly with [Co(terpy)2]2+, [Ru(NH3)5py]2+, and [Co(sep)]2+as reductants for horse cytochrome c(III). Previous work using CDNP-modified horse cytochrome c derivatives has indicated that the positively charged redox partners [Co(phen)3]3+(oxidant) and [Co(sep)]2+(reductant) react preferentially at site II on the front face of the molecule to the right-hand side of the solvent-accessible heme edge (the latter vertical with axially coordinated Met to the left), whereas negatively charged [Fe(CN)6]3-reacts at site III to the left-hand side and across the top section of the exposed heme edge. His33 is close to site II, consistent with the effect of protonation of this residue on reactivity with the 3+ oxidants. Reduction potentials for horse cytochrome c determined electrochemically in the presence of a mediator show no variation with pH over the range 7.5-5. Protonation of His33, the imidazole ring of which is 12 A from Fe, has no effect on the reduction potential therefore, and its effect on reactivity is a local effect. It is not immediately clear why there is no dependence on pH for the reaction of cytochrome c(III) with the three positively charged reductants. Possible contributing factors include some sort of conformational change, with a resultant shift in the site for electron transfer in the case of for example [Co(terpy)2]2+as compared to [Co(terpy)2]3+, and effects stemming from different degrees of solvation at or near His33. His33 of horse cytochrome c gives virtually identical pAa’s and rates of DEPC modification for both the oxidized and reduced forms, indicating similar degrees of accessibility.

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DO - 10.1021/ic00306a029

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SN - 0020-1669

VL - 28

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EP - 1366

JO - Inorganic Chemistry

JF - Inorganic Chemistry

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ER -

pH Dependence of Rate Constants for Reactions of Cytochrome c with Inorganic Redox Partners and Mechanistic Implications

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