12 Spectro-Electrochemical Investigation of the bc 1 Complex from the yeast Saccharomyces cerevisiae using Surface Enhanced B-Band Resonance Raman Spectroscopy !"#$%"&'()*() +,- ,&.*/(&0/12"/3%()4*56 +,- ,&7)/$%816)&91:*4 +,- ,&& ;1<=>*#>&?#1<< - &*#@&A"#*8"&BC&7C&&9*D5*## +,- & ! "#$%&'#()*%+(, /-0%123%&2'4503%60,0#3)78%9)*035#((:0;%!<8%==!>?%"#.(@8%%%% > 9/,-3.#(%+(, /-0%21%A0)7(2'2;4%B5CD%E%9+A8%F2(#/GH 4%I-3J%!8%!>><%K.0((#8%% ! B035#(4% > 9/,-3.#% 1. 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Materials and methods b<"(8/1()"5$(*<& 5"*%D/"5"#8%& KbM'& *#@& 7^L& :"/"& 6"/=1/5"@& 1#& 8"56<*8"& %8/$66"@& >1<@& KE'0L&"<"(8/1@"%,&:)"/"*%&618"#8$15"8/$(&8$8/*8$1#%&=1<<1:"@&FI&'bAA'&:"/"&(1#@D(8"@&1#& 8)"&816&6<*#"&%D/=*("%&1=&%$<]"/&/1@%&51@$=$"@&:$8)&O>9P%C& Modification of the silver electrodes :*%&@1#"&*%&@"%(/$F"@CK0/1%%"/D"%()4*56,&91:*4&"8& *<C&-WWSL&E)"&816&6<*#"%&1=&+-&55&8)$(4&%$<]"/&/1@%&:"/"&61<$%)"@&@1:#&81&*#&/5%&/1D>)#"%%& 1=&YH_& #5CK0/1%%"/D"%()4*56,& c/$"@/$()& "8& *<C& -WWSL& '"<=H*%%"5F<"@& 51#1<*I"/%& K'O.%L& 1=& (I%8"*5$#"&K7OL&:"/"&=1/5"@&1#&8)"&%$<]"/&/1@%&FI&$55"/%$1#&$#81&*#&*GD"1D%&%1<D8$1#&1=& 7O&K+W&5.f<L&=1/&+&)C&O=8"/&8)1/1D>)&/$#%$#>&:$8)&:*8"/&8)"&51@$=$"@&%$<]"/&%D/=*("%&:"/"& $55"/%"@& =1/& 8:1& )1D/%& $#81& *& %D%6"#%$1#& 1=& %$<]"/& #*#16*/8$(<"%& 1=& [W& #5& @$*5"8"/C& E)"/"*=8"/&8)"&%$<]"/&/1@%&:"/"&*>*$#&/$#%"@&:$8)&:*8"/C&& Immobilization of the protein and reconstitution into a lipid bilayer: &b$8)"/&E'0& "<"(8/1@"%& 1/& 8)"& *F1]"& 5"#8$1#"@& %$<]"/& "<"(8/1@"%& 51@$=$"@& :$8)& #*#16*/8$(<"%& :"/"& $55"/%"@&=1/&+&)&$#81&*&%1<D8$1#&1=&7O&+W&5.fBC&E)"&"J("%%&7O&:*%&/"51]"@&FI&/$#%$#>& 8)"& %D/=*("%& :$8)& :*8"/C& E)"& F( + &(156<"J&=/15&8)"&I"*%8&I#))7#3254)0,% )030L.,.#0&:*%& "J6/"%%"@&*#@&6D/$=$"@&*((1/@$#>&81&8)"&5"8)1@&1=&VC&Pg<%@h88$/&*#@&7C&VD#8"C(Pálsdóttir and Hunte 2003)&&E)"&6/18"$#&:*%&*@%1/F"@&FI&"J61%$#>&"$8)"/&8)"& >1<@& "<"(8/1@"& 1/&8)"&%$<]"/& %D/=*("%&=1/&+)&81&*&%1<D8$1#&1=&F( + &(156<"J&KWC+&i.f<L&$#&6)1%6)*8"&f&@1@"(I<H!H5*<81%$@"& K!!.L&FD=="/&K? 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Biochim Biophys Acta 416(2): 1 69- 89 Spiro, T G ( 197 8) "Resonance Raman spectra of hemoproteins." Methods Enzymol 54: 233 49 Spiro, T G ( 198 8) Biological applications of Raman spectroscopy, Vol 3: Resonance Raman spectra of heme proteins and other metalloproteins Spiro, T G., R S Czernuszewicz, et al ( 198 9) "Metalloporphyrin... swelling-deswelling changes in the gel (Fugure 1) Since being first reported in 199 6 as a “self-oscillating gel” (Yoshida et al., 199 6; 199 7), the author have been systematically studying the self-oscillating polymer and gel as well as their applications to biomimetic or smart materials (Figure 2) In fact, as applications to autonomic biomimetic actuators, ciliary motion actuators (Tabata et al., 2002, 2003)... in a biomimetic surface architecture." Biophys J 94 (9) : 3 698 -3705 Gao, F., H Qin, et al ( 199 8) "Q-band resonance Raman spectra of oxidized and reduced mitochondrial bc1 complexes." Biochemistry 37(27): 97 51-8 Grammel, H and R Ghosh (2008) "Redox-State Dynamics of Ubiquinone-10 Imply Cooperative Regulation of Photosynthetic Membrane Expression in Rhodospirillum rubrum." Journal of Bacteriology 190 , No.14:... Raman Spectroscopy frequency 343 356 635 650 663 681 697 716 747 783 92 0 1147 1170 1358 1374 1 390 1466 1 491 1502 15 59 1581 15 89 1606 1620 1631 mode assignment '8 (b H +c1 ), 8 (b H +c1 ) '50 (c1 ) '48 (c1 ) '20 (c1 ) '20 (c1 ) '7 (b), 7 (b) '7 (c1 ) '5 (b+c1 ) '15 (b+c1 ) '6 (c1 ) '46 (c1 ) '43 (c1 ) '30 (b H +c1 ), 30 (b H +c1 ) '4 (b+c1 ) 4 (b+c1 ) ' 29 (b+c1 ) '28 (b+c1 ) '3 (b+c1 ) 3 (b+c1 ) '38 (b+c1... Bacteriology 190 , No.14: 491 2- 492 1 Grosserueschkamp, M., M G Friedrich, et al (20 09) "Electron transfer kinetics of cytochrome c probed by time-resolved surface-enhanced resonance Raman spectroscopy." J Phys Chem B 113(8): 2 492 -7 Grosserueschkamp, M., C Nowak, et al (20 09) "Silver Surfaces with Optimized Surface Enhancement by Self-Assembly of Silver Nanoparticles for Spectroelectrochemical Applications. " Journal... ox red red red ox red ox red red red 263 265 377 403 407 416 681 / 696 96 8 '9 (b) '9 (b) 'CP r(b), 'CP r(b) 'CCaCb2,4 (c1 ) 'CCVn (b L ), CCVn (b L ) 'CCVn (b H ), CCVn (b H ) 'CS(c1 ), CS(c1 ) 'CP r(b+c1 ), CP r(b+c 1) 'CP r(b+c1 ), CP r(b+c1 ) 'CVn (b), CVn (b) ox red red red ox red red, ox red 97 2 1006 1087 1113 1130 1222 1308 14 29 'CCH3 (c1 ) '5 (b+c1 ) '14 (b+c1 ), 14 (b+c1 ) '13 (b), 13 (b) 'CHVn... cm-1 and at 14 29 cm-1 occur in the high frequency region, which originate from peripheral heme modes of the b hemes, CHVn and CH2Vn, respectively We found only one band at 12 69 cm-1 in the high frequency region, which was not considered in previous studies.(Le Moigne, Schoepp et al 199 9) In summary, the high frequency region contains bands, which originate from all types of hemes Of particular significance... the yeast cytochrome bc1 complex with its bound substrate cytochrome c." Proc Natl Acad Sci U S A 99 (5): 2800-5 Lange, C., J H Nett, et al (2001) "Specific roles of protein-phospholipid interactions in the yeast cytochrome bc1 complex structure." EMBO J 20(23): 6 591 -600 Le Moigne, C., B Schoepp, et al ( 199 9) "Distinct structures and environments for the three hemes of the cytochrome bc1 complex from... J.Solid State Electrochemistry DOI 10.1007/s10008-0101032-x Osawa, M., N Matsuda, et al ( 199 4) "Charge transfer resonance Raman process in surfaceenhanced Raman scattering from p-aminothiophenol adsorbed on silver: HerzbergTeller contribution." The Journal of Physical Chemistry 98 (48): 12702-12707 332 Biomimetic Based Applications Osyczka, A., C C Moser, et al (2005) "Fixing the Q cycle." TRENDS in Biochemical . cm -1 15 89 cm -1 1581 cm -1 15 59 cm -1 1502 cm -1 1 491 cm -1 1466 cm -1 14 29 cm -1 1 390 cm -1 1374 cm -1 1358 cm -1 1308 cm -1 12 69 cm -1 intensity / a.u. 'Q /cm -1 C D 850 90 0 95 0 1000. cm -1 697 cm -1 681 cm -1 663 cm -1 650 cm -1 635 cm -1 602 cm -1 560 cm -1 503 cm -1 562 cm -1 'Q /cm -1 546 cm -1 518 cm -1 460 cm -1 a A B & & % Biomimetic Based Applications. /-0%123%&2'4503%60,0#3)78 %9) *035#((:0;%!<8%==!>?%"#.(@8%%%% > 9/ ,-3.#(%+(, /-0%21%A0)7(2'2;4%B5CD%E %9+ A8%F2(#/GH 4%I-3J%!8%!>><%K.0((#8%% ! B035#(4% > 9/ ,-3.#% 1. Introduction