Investigation of the electron-transfer mechanism by cross-linking between Zn-substituted myoglobin and cytochrome b5

Yoshiaki Furukawa, Fumihiro Matsuda, Koichiro Ishimori, Isao Morishima

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

We have investigated the photoinduced electron transfer (ET) in the 1:1 cross-linked complex (CL-ZnMb/b5) formed by a cross-linking reagent, EDC, between Zn-substituted myoglobin (ZnMb) and cytochrome b5 (Cytb5) to reveal the mechanism of the inter-protein ET reactions under the condition of multiple encounter complexes. A variety of the ZnMb-Cytb5 orientations was suggested because of failure to identify the single and specific cross-linking site on Cytb5 by the peptide-mapping analysis using mass spectrometry. In CL-ZnMb/b5, a laser pulse generates the triplet excited state of the ZnMb domain (3ZnMb*), which can transfer one electron to the Cytb5 domain. The decay kinetics of 3ZnMb* in CL-ZnMb/ b5 consists of a facile power-law ET phase to Cytb5 domain (∼30%) and a slower single-exponential phase (∼70%). The application of the Marcus equation to this power-law phase indicates that CL-ZnMb/b5 has a variety of ZnMb-Cytb5 orientations for the facile ET in which the distance between the redox centers (D-A distance) is distributed over 13-20 Å. The single-exponential phase in the 3ZnMb* decay kinetics of CL ZnMb/b5 is similar to the intrinsic decay of 3ZnMb* in its rate constant, 65 s-1. This implies that the ET is impeded in about 70% of the total ZnMb-Cytb5 orientations due to the D-A distance larger than 20 Å. Combined with the results of the Brownian dynamics simulations for the encounter complexes, the overall bimolecular ET rate, kapp, can be reproduced by the sum of the ET rates for the minor encounter complexes of which D-A distance is less than 20 Å. On the other hand, the encounter complexes with longer D-A distance, which are the majority of the encounter complexes between ZnMb and Cytb5, have little contribution to the overall bimolecular ET rate. These observations experimentally demonstrate that ZnMb forms a variety of encounter complexes with Cytb5, among which a minor set of the complexes with the shorter D-A distance (<∼20 Å) regulates the overall bimolecular ET between the proteins.

Original languageEnglish
Pages (from-to)4008-4019
Number of pages12
JournalJournal of the American Chemical Society
Volume124
Issue number15
DOIs
Publication statusPublished - 2002 Apr 17
Externally publishedYes

Fingerprint

Cytochromes b5
Myoglobin
Electrons
Proteins
Cross-Linking Reagents
Kinetics
Peptide Mapping
Excited states
Peptides
Mass spectrometry
Laser pulses
Rate constants
Oxidation-Reduction
Mass Spectrometry
Lasers

ASJC Scopus subject areas

  • Chemistry(all)

Cite this

Investigation of the electron-transfer mechanism by cross-linking between Zn-substituted myoglobin and cytochrome b5 . / Furukawa, Yoshiaki; Matsuda, Fumihiro; Ishimori, Koichiro; Morishima, Isao.

In: Journal of the American Chemical Society, Vol. 124, No. 15, 17.04.2002, p. 4008-4019.

Research output: Contribution to journalArticle

@article{31e859fc7fcf426b937597b98edc3ddc,
title = "Investigation of the electron-transfer mechanism by cross-linking between Zn-substituted myoglobin and cytochrome b5",
abstract = "We have investigated the photoinduced electron transfer (ET) in the 1:1 cross-linked complex (CL-ZnMb/b5) formed by a cross-linking reagent, EDC, between Zn-substituted myoglobin (ZnMb) and cytochrome b5 (Cytb5) to reveal the mechanism of the inter-protein ET reactions under the condition of multiple encounter complexes. A variety of the ZnMb-Cytb5 orientations was suggested because of failure to identify the single and specific cross-linking site on Cytb5 by the peptide-mapping analysis using mass spectrometry. In CL-ZnMb/b5, a laser pulse generates the triplet excited state of the ZnMb domain (3ZnMb*), which can transfer one electron to the Cytb5 domain. The decay kinetics of 3ZnMb* in CL-ZnMb/ b5 consists of a facile power-law ET phase to Cytb5 domain (∼30{\%}) and a slower single-exponential phase (∼70{\%}). The application of the Marcus equation to this power-law phase indicates that CL-ZnMb/b5 has a variety of ZnMb-Cytb5 orientations for the facile ET in which the distance between the redox centers (D-A distance) is distributed over 13-20 {\AA}. The single-exponential phase in the 3ZnMb* decay kinetics of CL ZnMb/b5 is similar to the intrinsic decay of 3ZnMb* in its rate constant, 65 s-1. This implies that the ET is impeded in about 70{\%} of the total ZnMb-Cytb5 orientations due to the D-A distance larger than 20 {\AA}. Combined with the results of the Brownian dynamics simulations for the encounter complexes, the overall bimolecular ET rate, kapp, can be reproduced by the sum of the ET rates for the minor encounter complexes of which D-A distance is less than 20 {\AA}. On the other hand, the encounter complexes with longer D-A distance, which are the majority of the encounter complexes between ZnMb and Cytb5, have little contribution to the overall bimolecular ET rate. These observations experimentally demonstrate that ZnMb forms a variety of encounter complexes with Cytb5, among which a minor set of the complexes with the shorter D-A distance (<∼20 {\AA}) regulates the overall bimolecular ET between the proteins.",
author = "Yoshiaki Furukawa and Fumihiro Matsuda and Koichiro Ishimori and Isao Morishima",
year = "2002",
month = "4",
day = "17",
doi = "10.1021/ja0171916",
language = "English",
volume = "124",
pages = "4008--4019",
journal = "Journal of the American Chemical Society",
issn = "0002-7863",
publisher = "American Chemical Society",
number = "15",

}

TY - JOUR

T1 - Investigation of the electron-transfer mechanism by cross-linking between Zn-substituted myoglobin and cytochrome b5

AU - Furukawa, Yoshiaki

AU - Matsuda, Fumihiro

AU - Ishimori, Koichiro

AU - Morishima, Isao

PY - 2002/4/17

Y1 - 2002/4/17

N2 - We have investigated the photoinduced electron transfer (ET) in the 1:1 cross-linked complex (CL-ZnMb/b5) formed by a cross-linking reagent, EDC, between Zn-substituted myoglobin (ZnMb) and cytochrome b5 (Cytb5) to reveal the mechanism of the inter-protein ET reactions under the condition of multiple encounter complexes. A variety of the ZnMb-Cytb5 orientations was suggested because of failure to identify the single and specific cross-linking site on Cytb5 by the peptide-mapping analysis using mass spectrometry. In CL-ZnMb/b5, a laser pulse generates the triplet excited state of the ZnMb domain (3ZnMb*), which can transfer one electron to the Cytb5 domain. The decay kinetics of 3ZnMb* in CL-ZnMb/ b5 consists of a facile power-law ET phase to Cytb5 domain (∼30%) and a slower single-exponential phase (∼70%). The application of the Marcus equation to this power-law phase indicates that CL-ZnMb/b5 has a variety of ZnMb-Cytb5 orientations for the facile ET in which the distance between the redox centers (D-A distance) is distributed over 13-20 Å. The single-exponential phase in the 3ZnMb* decay kinetics of CL ZnMb/b5 is similar to the intrinsic decay of 3ZnMb* in its rate constant, 65 s-1. This implies that the ET is impeded in about 70% of the total ZnMb-Cytb5 orientations due to the D-A distance larger than 20 Å. Combined with the results of the Brownian dynamics simulations for the encounter complexes, the overall bimolecular ET rate, kapp, can be reproduced by the sum of the ET rates for the minor encounter complexes of which D-A distance is less than 20 Å. On the other hand, the encounter complexes with longer D-A distance, which are the majority of the encounter complexes between ZnMb and Cytb5, have little contribution to the overall bimolecular ET rate. These observations experimentally demonstrate that ZnMb forms a variety of encounter complexes with Cytb5, among which a minor set of the complexes with the shorter D-A distance (<∼20 Å) regulates the overall bimolecular ET between the proteins.

AB - We have investigated the photoinduced electron transfer (ET) in the 1:1 cross-linked complex (CL-ZnMb/b5) formed by a cross-linking reagent, EDC, between Zn-substituted myoglobin (ZnMb) and cytochrome b5 (Cytb5) to reveal the mechanism of the inter-protein ET reactions under the condition of multiple encounter complexes. A variety of the ZnMb-Cytb5 orientations was suggested because of failure to identify the single and specific cross-linking site on Cytb5 by the peptide-mapping analysis using mass spectrometry. In CL-ZnMb/b5, a laser pulse generates the triplet excited state of the ZnMb domain (3ZnMb*), which can transfer one electron to the Cytb5 domain. The decay kinetics of 3ZnMb* in CL-ZnMb/ b5 consists of a facile power-law ET phase to Cytb5 domain (∼30%) and a slower single-exponential phase (∼70%). The application of the Marcus equation to this power-law phase indicates that CL-ZnMb/b5 has a variety of ZnMb-Cytb5 orientations for the facile ET in which the distance between the redox centers (D-A distance) is distributed over 13-20 Å. The single-exponential phase in the 3ZnMb* decay kinetics of CL ZnMb/b5 is similar to the intrinsic decay of 3ZnMb* in its rate constant, 65 s-1. This implies that the ET is impeded in about 70% of the total ZnMb-Cytb5 orientations due to the D-A distance larger than 20 Å. Combined with the results of the Brownian dynamics simulations for the encounter complexes, the overall bimolecular ET rate, kapp, can be reproduced by the sum of the ET rates for the minor encounter complexes of which D-A distance is less than 20 Å. On the other hand, the encounter complexes with longer D-A distance, which are the majority of the encounter complexes between ZnMb and Cytb5, have little contribution to the overall bimolecular ET rate. These observations experimentally demonstrate that ZnMb forms a variety of encounter complexes with Cytb5, among which a minor set of the complexes with the shorter D-A distance (<∼20 Å) regulates the overall bimolecular ET between the proteins.

UR - http://www.scopus.com/inward/record.url?scp=0037123265&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0037123265&partnerID=8YFLogxK

U2 - 10.1021/ja0171916

DO - 10.1021/ja0171916

M3 - Article

VL - 124

SP - 4008

EP - 4019

JO - Journal of the American Chemical Society

JF - Journal of the American Chemical Society

SN - 0002-7863

IS - 15

ER -