Role of methionine 230 in intramolecular electron transfer between the oxyferryl heme and tryptophan 191 in cytochrome c peroxidase compound II

Peroxide-activated Auricularia auricula-judae dye-decolorizing peroxidase (DyP) forms a mixed Trp377 and Tyr337 radical, the former being responsible for oxidation of the typical DyP substrates (Linde et al. Biochem. J., 2015, 466, 253-262); however, a pure tryptophanyl radical EPR signal is detected at pH 7 (where the enzyme is inactive), in contrast with the mixed signal observed at pH for optimum activity, pH 3. On the contrary, the presence of a second tyrosine radical (at Tyr147) is deduced by a multifrequency EPR study of a variety of simple and double-directed variants (including substitution of the above and other tryptophan and tyrosine residues) at different freezing times after their activation by H2O2 (at pH 3). This points out that subsidiary long-range electron-transfer pathways enter into operation when the main pathway(s) is removed by directed mutagenesis, with catalytic efficiencies progressively decreasing. Finally, self-reduction of the Trp377 neutral radical is observed when reaction time (before freezing) is increased in the absence of reducing substrates (from 10 to 60 s). Interestingly, the tryptophanyl radical is stable in the Y147S/Y337S variant, indicating that these two tyrosine residues are involved in the self-reduction reaction.

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“…It has been shown that methionine residues play a role in the electrostatic stabilization of Trp protein radicals 47,25,62,[66][67][68] .…”

Section: Lret Pathway/s In Aaudypmentioning

confidence: 99%

Redox-Active Sites in Auricularia auricula-judae Dye-Decolorizing Peroxidase and Several Directed Variants: A Multifrequency EPR Study

et al. 2015

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“…However, this simple picture became clouded with more detailed studies and it was found that with yeast cytc the activity actually increases with increasing ionic strength and only at moderately high salt concentrations does the activity decrease [77]. The reason for such behavior may be related to a change in rate limiting step as a function of ionic strength [56] and to best understand this explanation we refer to the following scheme fashioned after [78]. …”

Section: Electron Transfermentioning

confidence: 99%

Thirty years of heme peroxidase structural biology

Poulos

2010

Archives of Biochemistry and Biophysics

108

106

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“…All of the Ru-Cc derivatives were found to initially reduce the Trp-191 radical cation in CMPI(Fe IV d O,R •+ ) as shown in Scheme 1 (18,19). CMPII(Fe IV dO,R) is then converted to CMPII(Fe III ,R •+ ) by intramolecular electron transfer, followed by reduction of the radical cation in CMPII(Fe III ,R •+ ) by a second molecule of Cc II which gives the resting enzyme (21). Stopped-flow studies have demonstrated that native hCc and yCc initially reduce the radical cation in CMPI according to Scheme 1 under all conditions of ionic strength and pH (22,24,25).…”

mentioning

confidence: 99%

Role of Configurational Gating in Intracomplex Electron Transfer from Cytochrome c to the Radical Cation in Cytochrome c Peroxidase

et al. 1999

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Electron transfer within complexes of cytochrome c (Cc) and cytochrome c peroxidase (CcP) was studied to determine whether the reactions are gated by fluctuations in configuration. Electron transfer in the physiological complex of yeast Cc (yCc) and CcP was studied using the Ru-39-Cc derivative, in which the H39C/C102T variant of yeast iso-1-cytochrome c is labeled at the single cysteine residue on the back surface with trisbipyridylruthenium(II). Laser excitation of the 1:1 Ru-39-Cc-CcP compound I complex at low ionic strength results in rapid electron transfer from RuII to heme c FeIII, followed by electron transfer from heme c FeII to the Trp-191 indolyl radical cation with a rate constant keta of 2 x 10(6) s-1 at 20 degrees C. keta is not changed by increasing the viscosity up to 40 cP with glycerol and is independent of temperature. These results suggest that this reaction is not gated by fluctuations in the configuration of the complex, but may represent the elementary electron transfer step. The value of keta is consistent with the efficient pathway for electron transfer in the crystalline yCc-CcP complex, which has a distance of 16 A between the edge of heme c and the Trp-191 indole [Pelletier, H., and Kraut, J. (1992) Science 258, 1748-1755]. Electron transfer in the complex of horse Cc (hCc) and CcP was examined using Ru-27-Cc, in which hCc is labeled with trisbipyridylruthenium(II) at Lys-27. Laser excitation of the Ru-27-Cc-CcP complex results in electron transfer from RuII to heme c FeII with a rate constant k1 of 2.3 x 10(7) s-1, followed by oxidation of the Trp-191 indole to a radical cation by RuIII with a rate constant k3 of 7 x 10(6) s-1. The cycle is completed by electron transfer from heme c FeII to the Trp-191 radical cation with a rate constant k4 of 6.1 x 10(4) s-1. The rate constant k4 decreases to 3.4 x 10(3) s-1 as the viscosity is increased to 84 cP, but the rate constants k1 and k3 remain the same. The results are consistent with a gating mechanism in which the Ru-27-Cc-CcP complex undergoes fluctuations between a major state A with the configuration of the hCc-CcP crystalline complex and a minor state B with the configuration of the yCc-CcP complex. The hCc-CcP complex, state A, has an inefficient pathway for electron transfer from heme c to the Trp-191 indolyl radical cation with a distance of 20.5 A and a predicted value of 5 x 10(2) s-1 for k4A. The observed rate constant k4 is thus gated by the rate constant ka for conversion of state A to state B, where the rate of electron transfer k4B is expected to be 2 x 10(6) s-1. The temperature dependence of k4 provides activation parameters that are consistent with the proposed gating mechanism. These studies provide evidence that configurational gating does not control electron transfer in the physiological yCc-CcP complex, but is required in the nonphysiological hCc-CcP complex.

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Role of methionine 230 in intramolecular electron transfer between the oxyferryl heme and tryptophan 191 in cytochrome c peroxidase compound II

Cited by 49 publications

References 33 publications

Redox-Active Sites in Auricularia auricula-judae Dye-Decolorizing Peroxidase and Several Directed Variants: A Multifrequency EPR Study

Redox-Active Sites in Auricularia auricula-judae Dye-Decolorizing Peroxidase and Several Directed Variants: A Multifrequency EPR Study

Thirty years of heme peroxidase structural biology

Role of Configurational Gating in Intracomplex Electron Transfer from Cytochrome c to the Radical Cation in Cytochrome c Peroxidase

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