Evidence for concerted electron proton transfer in charge recombination between FADH- and 306Trp in Escherichia coli photolyase

Agnieszka A. Zieba, Caroline Richardson, Carlos Lucero, Senghane D. Dieng, Yvonne Gindt, Johannes Schelvis

Research output: Contribution to journalArticleResearchpeer-review

15 Citations (Scopus)

Abstract

Proton-coupled electron-transfer (PCET) is a mechanism of great importance in protein electron transfer and enzyme catalysis, and the involvement of aromatic amino acids in this process is of much interest. The DNA repair enzyme photolyase provides a natural system that allows for the study of PCET using a neutral radical tryptophan (Trp). In Escherichia coli photolyase, photoreduction of the flavin adenine dinucleotide (FAD) cofactor in its neutral radical semiquinone form (FADH) results in the formation of FADH- and 306Trp. Charge recombination between these two intermediates requires the uptake of a proton by 306Trp. The rate constant of charge recombination has been measured as a function of temperature in the pH range from 5.5 to 10.0, and the data are analyzed with both classical Marcus and semi-classical Hopfield electron transfer theory. The reorganization energy associated with the charge recombination process shows a pH dependence ranging from 2.3 eV at pH ? 7 and 1.2 eV at pH(D) 10.0. These findings indicate that at least two mechanisms are involved in the charge recombination reaction. Global analysis of the data supports the hypothesis that PCET during charge recombination can follow two different mechanisms with an apparent switch around pH 6.5. At lower pH, concerted electron proton transfer (CEPT) is the favorable mechanism with a reorganization energy of 2.1-2.3 eV. At higher pH, a sequential mechanism becomes dominant with rate-limiting electron-transfer followed by proton uptake which has a reorganization energy of 1.0-1.3 eV. The observed ?inverse? deuterium isotope effect at pH < 8 can be explained by a solvent isotope effect that affects the free energy change of the reaction and masks the normal, mass-related kinetic isotope effect that is expected for a CEPT mechanism. To the best of our knowledge, this is the first time that a switch in PCET mechanism has been observed in a protein.

Original languageEnglish
Pages (from-to)7824-7836
Number of pages13
JournalJournal of the American Chemical Society
Volume133
Issue number20
DOIs
StatePublished - 25 May 2011

Fingerprint

Deoxyribodipyrimidine Photo-Lyase
Proton transfer
Escherichia coli
Genetic Recombination
Protons
Electrons
Isotopes
Switches
DNA Repair Enzymes
Proteins
Aromatic Amino Acids
Flavin-Adenine Dinucleotide
Deuterium
Masks
Catalysis
Carboxylic acids
Tryptophan
Free energy
Charge transfer
Amino acids

Cite this

@article{9d0b8426299248c3b5b1e0d60adaa63c,
title = "Evidence for concerted electron proton transfer in charge recombination between FADH- and 306Trp• in Escherichia coli photolyase",
abstract = "Proton-coupled electron-transfer (PCET) is a mechanism of great importance in protein electron transfer and enzyme catalysis, and the involvement of aromatic amino acids in this process is of much interest. The DNA repair enzyme photolyase provides a natural system that allows for the study of PCET using a neutral radical tryptophan (Trp•). In Escherichia coli photolyase, photoreduction of the flavin adenine dinucleotide (FAD) cofactor in its neutral radical semiquinone form (FADH•) results in the formation of FADH- and 306Trp•. Charge recombination between these two intermediates requires the uptake of a proton by 306Trp•. The rate constant of charge recombination has been measured as a function of temperature in the pH range from 5.5 to 10.0, and the data are analyzed with both classical Marcus and semi-classical Hopfield electron transfer theory. The reorganization energy associated with the charge recombination process shows a pH dependence ranging from 2.3 eV at pH ? 7 and 1.2 eV at pH(D) 10.0. These findings indicate that at least two mechanisms are involved in the charge recombination reaction. Global analysis of the data supports the hypothesis that PCET during charge recombination can follow two different mechanisms with an apparent switch around pH 6.5. At lower pH, concerted electron proton transfer (CEPT) is the favorable mechanism with a reorganization energy of 2.1-2.3 eV. At higher pH, a sequential mechanism becomes dominant with rate-limiting electron-transfer followed by proton uptake which has a reorganization energy of 1.0-1.3 eV. The observed ?inverse? deuterium isotope effect at pH < 8 can be explained by a solvent isotope effect that affects the free energy change of the reaction and masks the normal, mass-related kinetic isotope effect that is expected for a CEPT mechanism. To the best of our knowledge, this is the first time that a switch in PCET mechanism has been observed in a protein.",
author = "Zieba, {Agnieszka A.} and Caroline Richardson and Carlos Lucero and Dieng, {Senghane D.} and Yvonne Gindt and Johannes Schelvis",
year = "2011",
month = "5",
day = "25",
doi = "10.1021/ja2001488",
language = "English",
volume = "133",
pages = "7824--7836",
journal = "Journal of the American Chemical Society",
issn = "0002-7863",
publisher = "American Chemical Society",
number = "20",

}

Evidence for concerted electron proton transfer in charge recombination between FADH- and 306Trp in Escherichia coli photolyase. / Zieba, Agnieszka A.; Richardson, Caroline; Lucero, Carlos; Dieng, Senghane D.; Gindt, Yvonne; Schelvis, Johannes.

In: Journal of the American Chemical Society, Vol. 133, No. 20, 25.05.2011, p. 7824-7836.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Evidence for concerted electron proton transfer in charge recombination between FADH- and 306Trp• in Escherichia coli photolyase

AU - Zieba, Agnieszka A.

AU - Richardson, Caroline

AU - Lucero, Carlos

AU - Dieng, Senghane D.

AU - Gindt, Yvonne

AU - Schelvis, Johannes

PY - 2011/5/25

Y1 - 2011/5/25

N2 - Proton-coupled electron-transfer (PCET) is a mechanism of great importance in protein electron transfer and enzyme catalysis, and the involvement of aromatic amino acids in this process is of much interest. The DNA repair enzyme photolyase provides a natural system that allows for the study of PCET using a neutral radical tryptophan (Trp•). In Escherichia coli photolyase, photoreduction of the flavin adenine dinucleotide (FAD) cofactor in its neutral radical semiquinone form (FADH•) results in the formation of FADH- and 306Trp•. Charge recombination between these two intermediates requires the uptake of a proton by 306Trp•. The rate constant of charge recombination has been measured as a function of temperature in the pH range from 5.5 to 10.0, and the data are analyzed with both classical Marcus and semi-classical Hopfield electron transfer theory. The reorganization energy associated with the charge recombination process shows a pH dependence ranging from 2.3 eV at pH ? 7 and 1.2 eV at pH(D) 10.0. These findings indicate that at least two mechanisms are involved in the charge recombination reaction. Global analysis of the data supports the hypothesis that PCET during charge recombination can follow two different mechanisms with an apparent switch around pH 6.5. At lower pH, concerted electron proton transfer (CEPT) is the favorable mechanism with a reorganization energy of 2.1-2.3 eV. At higher pH, a sequential mechanism becomes dominant with rate-limiting electron-transfer followed by proton uptake which has a reorganization energy of 1.0-1.3 eV. The observed ?inverse? deuterium isotope effect at pH < 8 can be explained by a solvent isotope effect that affects the free energy change of the reaction and masks the normal, mass-related kinetic isotope effect that is expected for a CEPT mechanism. To the best of our knowledge, this is the first time that a switch in PCET mechanism has been observed in a protein.

AB - Proton-coupled electron-transfer (PCET) is a mechanism of great importance in protein electron transfer and enzyme catalysis, and the involvement of aromatic amino acids in this process is of much interest. The DNA repair enzyme photolyase provides a natural system that allows for the study of PCET using a neutral radical tryptophan (Trp•). In Escherichia coli photolyase, photoreduction of the flavin adenine dinucleotide (FAD) cofactor in its neutral radical semiquinone form (FADH•) results in the formation of FADH- and 306Trp•. Charge recombination between these two intermediates requires the uptake of a proton by 306Trp•. The rate constant of charge recombination has been measured as a function of temperature in the pH range from 5.5 to 10.0, and the data are analyzed with both classical Marcus and semi-classical Hopfield electron transfer theory. The reorganization energy associated with the charge recombination process shows a pH dependence ranging from 2.3 eV at pH ? 7 and 1.2 eV at pH(D) 10.0. These findings indicate that at least two mechanisms are involved in the charge recombination reaction. Global analysis of the data supports the hypothesis that PCET during charge recombination can follow two different mechanisms with an apparent switch around pH 6.5. At lower pH, concerted electron proton transfer (CEPT) is the favorable mechanism with a reorganization energy of 2.1-2.3 eV. At higher pH, a sequential mechanism becomes dominant with rate-limiting electron-transfer followed by proton uptake which has a reorganization energy of 1.0-1.3 eV. The observed ?inverse? deuterium isotope effect at pH < 8 can be explained by a solvent isotope effect that affects the free energy change of the reaction and masks the normal, mass-related kinetic isotope effect that is expected for a CEPT mechanism. To the best of our knowledge, this is the first time that a switch in PCET mechanism has been observed in a protein.

UR - http://www.scopus.com/inward/record.url?scp=79957701723&partnerID=8YFLogxK

U2 - 10.1021/ja2001488

DO - 10.1021/ja2001488

M3 - Article

VL - 133

SP - 7824

EP - 7836

JO - Journal of the American Chemical Society

JF - Journal of the American Chemical Society

SN - 0002-7863

IS - 20

ER -