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Title: Plasmon waveguide resonance spectroscopic evidence fordifferential binding of oxidized and reduced rhodobacter capsulatuscytochrome c(2) to the cytochrome bc(1) complex mediated by theconformation of the rieske iron-sulfur protein

Abstract

The dissociation constants for the binding of Rhodobactercapsulatus cytochrome c2 and its K93P mutant to the cytochrome bc1complex embedded in a phospholipid bilayer were measured by plasmonwaveguide resonance spectroscopy in the presence and absence of theinhibitor stigmatellin. The reduced form of cytochrome c2 strongly bindsto reduced cytochrome bc1 (Kd = 0.02 M) but binds much more weakly to theoxidized form (Kd = 3.1 M). In contrast, oxidized cytochrome c2 binds tooxidized cytochrome bc1 in a biphasic fashion with Kd values of 0.11 and0.58 M. Such a biphasic interaction is consistent with binding to twoseparate sites or conformations of oxidized cytochrome c2 and/orcytochrome bc1. However, in the presence of stigmatellin, we find thatoxidized cytochrome c2 binds to oxidized cytochrome bc1 in a monophasicfashion with high affinity (Kd = 0.06 M) and reduced cytochrome c2 bindsless strongly (Kd = 0.11 M) but ~;30-fold more tightly than in theabsence of stigmatellin. Structural studies with cytochrome bc1, with andwithout the inhibitor stigmatellin, have led to the proposal that theRieske protein is mobile, moving between the cytochrome b and cytochromec1 components during turnover. In one conformation, the Rieske proteinbinds near the heme of cytochrome c1, while the cytochrome c2 bindingsite is also near the cytochromemore » c1 heme but on the opposite side fromthe Rieske site, where cytochrome c2 cannot directly interact withRieske. However, the inhibitor, stigmatellin, freezes the Rieske proteiniron-sulfur cluster in a conformation proximal to cytochrome b and distalto cytochrome c1. We conclude from this that the dual conformation of theRieske protein is primarily responsible for biphasic binding of oxidizedcytochrome c2 to cytochrome c1. This optimizes turnover by maximizingbinding of the substrate, oxidized cytochrome c2, when the iron-sulfurcluster is proximal to cytochrome b and minimizing binding of theproduct, reduced cytochrome c2, when it is proximal to cytochromec1.« less

Authors:
; ; ; ; ; ;
Publication Date:
Research Org.:
COLLABORATION - University ofArizona
OSTI Identifier:
929360
Report Number(s):
LBNL-62862
R&D Project: 864U1F; TRN: US0803724
DOE Contract Number:  
DE-AC02-05CH11231
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemistry; Journal Volume: 46; Journal Issue: 24; Related Information: Journal Publication Date: Jun 19 2007
Country of Publication:
United States
Language:
English
Subject:
59; AFFINITY; CYTOCHROMES; DISSOCIATION; HEME; MUTANTS; PHOSPHOLIPIDS; PLASMONS; PROTEINS; RESONANCE; SPECTROSCOPY; WAVEGUIDES; plasmon waveguide resonance; photosynthetic reaction center; cytochrome bc1 complex; cytochrome c2; Rieske iron-sulfur protein; quinone; cytochrome b.

Citation Formats

Devanathan, S., Salamon, Z., Tollin, G., Fitch, J.C., Meyer,T.E., Berry, E.A., and Cusanovich, M.A. Plasmon waveguide resonance spectroscopic evidence fordifferential binding of oxidized and reduced rhodobacter capsulatuscytochrome c(2) to the cytochrome bc(1) complex mediated by theconformation of the rieske iron-sulfur protein. United States: N. p., 2007. Web. doi:10.1021/bi602649u.
Devanathan, S., Salamon, Z., Tollin, G., Fitch, J.C., Meyer,T.E., Berry, E.A., & Cusanovich, M.A. Plasmon waveguide resonance spectroscopic evidence fordifferential binding of oxidized and reduced rhodobacter capsulatuscytochrome c(2) to the cytochrome bc(1) complex mediated by theconformation of the rieske iron-sulfur protein. United States. doi:10.1021/bi602649u.
Devanathan, S., Salamon, Z., Tollin, G., Fitch, J.C., Meyer,T.E., Berry, E.A., and Cusanovich, M.A. Tue . "Plasmon waveguide resonance spectroscopic evidence fordifferential binding of oxidized and reduced rhodobacter capsulatuscytochrome c(2) to the cytochrome bc(1) complex mediated by theconformation of the rieske iron-sulfur protein". United States. doi:10.1021/bi602649u.
@article{osti_929360,
title = {Plasmon waveguide resonance spectroscopic evidence fordifferential binding of oxidized and reduced rhodobacter capsulatuscytochrome c(2) to the cytochrome bc(1) complex mediated by theconformation of the rieske iron-sulfur protein},
author = {Devanathan, S. and Salamon, Z. and Tollin, G. and Fitch, J.C. and Meyer,T.E. and Berry, E.A. and Cusanovich, M.A.},
abstractNote = {The dissociation constants for the binding of Rhodobactercapsulatus cytochrome c2 and its K93P mutant to the cytochrome bc1complex embedded in a phospholipid bilayer were measured by plasmonwaveguide resonance spectroscopy in the presence and absence of theinhibitor stigmatellin. The reduced form of cytochrome c2 strongly bindsto reduced cytochrome bc1 (Kd = 0.02 M) but binds much more weakly to theoxidized form (Kd = 3.1 M). In contrast, oxidized cytochrome c2 binds tooxidized cytochrome bc1 in a biphasic fashion with Kd values of 0.11 and0.58 M. Such a biphasic interaction is consistent with binding to twoseparate sites or conformations of oxidized cytochrome c2 and/orcytochrome bc1. However, in the presence of stigmatellin, we find thatoxidized cytochrome c2 binds to oxidized cytochrome bc1 in a monophasicfashion with high affinity (Kd = 0.06 M) and reduced cytochrome c2 bindsless strongly (Kd = 0.11 M) but ~;30-fold more tightly than in theabsence of stigmatellin. Structural studies with cytochrome bc1, with andwithout the inhibitor stigmatellin, have led to the proposal that theRieske protein is mobile, moving between the cytochrome b and cytochromec1 components during turnover. In one conformation, the Rieske proteinbinds near the heme of cytochrome c1, while the cytochrome c2 bindingsite is also near the cytochrome c1 heme but on the opposite side fromthe Rieske site, where cytochrome c2 cannot directly interact withRieske. However, the inhibitor, stigmatellin, freezes the Rieske proteiniron-sulfur cluster in a conformation proximal to cytochrome b and distalto cytochrome c1. We conclude from this that the dual conformation of theRieske protein is primarily responsible for biphasic binding of oxidizedcytochrome c2 to cytochrome c1. This optimizes turnover by maximizingbinding of the substrate, oxidized cytochrome c2, when the iron-sulfurcluster is proximal to cytochrome b and minimizing binding of theproduct, reduced cytochrome c2, when it is proximal to cytochromec1.},
doi = {10.1021/bi602649u},
journal = {Biochemistry},
number = 24,
volume = 46,
place = {United States},
year = {Tue May 22 00:00:00 EDT 2007},
month = {Tue May 22 00:00:00 EDT 2007}
}