IS-

Surface Plasmon Enhanced Interracial Electron Transfer and Resonance Raman, Surface-enhanced Resonance Raman Studies of Cytochrome C Mutants

by

Zheng, Junwe i

PHD Thesis submitted to Iowa State University

Ames Laboratory, U.S. DOE

Iowa State University

Ames, Iowa 50011

Date Transmitted: November 8, 1999

PREPARED FOR THE U.S. DEPARTMENT OF ENERGY

UNDER CONTRACT NO. W-7405-Eng-82.

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TABLE OF CONTENTS

CHAPTER 1. GENEIL4L INTRODUCTION Dissertation Organization The Mechanisms for Sutiace-enhanced Raman Scattering

Electromagnetic Enhancement Model Chemical Effect Model New Trends in Application of SERS

Cytochrome c and its Raman Scattering Spectroscopy Cytochrome c Resonance Raman Surface-enhanced Resonance Raman Scattering

References

CHAPTER 2. PHOTOINDUCED ELECTROCHEMICAL REDUCTION OF NITRITE AT ELECTROCHEIVDCALLY ROUGHENED SILVER SURFACE

Abstract Introduction Experimental Methods Results and Discussion

Effect of Electrode Material and Surface Treatment Effect of Excitation Wavelength on Photoelectrochemical Response Effect of pH Reduction of Nitrate

Conclusions Acknowledgement References

CHAPTER 3. PHOTOELECTROCHEMICAL REDUCTION OF COZMEDIATED WITH METHYLVIOLOGEN AT ROUGHENED SILVER ELECTRODES

Abstract Introduction Experimental

Chemicals Apparatus and Methods

Results Discussion Conclusion Acknowledgement References

1 2 3 4 6 7 8 8

17 23 23

29 29 29 32 34

34 34 49 51 51 53 53

56 56 56 58 58 58 60 72 76 76 76

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CHAPTER 4. PHOTOINDUCED ELECTRON TRANSFER AT THE SURFACE OF NANOSIZE SILVER PARTICLES AS MONITORED BY EPR SPECTROSCOPY 79

Abstract 79

Introduction 79

Experimental Methods 80

Results and Discussion 81

References 88

CHAPTER 5. ELECTROCHEMISTRY AND SURFACE-ENHANCED RESONANCE RAMAN SCATTERING SPECTRA OF MICROPEROXIDASE-1 1

Abstract Introduction Experimental Section

Materials and Solutions Apparatus and Procedures

Results and Discussion Electrochemistry of NIP-l 1 RR spectra of NW-l 1 SERRS SpectraofMP-11 Photo-induced Reduction ofMP-11

Conclusion Acknowledgement References

CHAPTER 6. RESONANCE RAMAN STUDY OF CYTOCHROME C WATER MUTANTS

Abstract Introduction Experimental Results and Discussion

Tyrosine-67 to phenylalanine Asparagine-52 to isoleucine Histidine-26 to valine

Conclusions Acknowledgement References

90 90 90 92 92 92 93 93 96 96

102 105 106 106

108 108 108 113 114 122 122 124 126 128 128

CHAPTER 7. CIRCULAR DICHROISM AND RESONANCE RAMAN COMPARA- TIVE STUDIES OF WILD TYPE CYTOCHROME C W F82H MUTANT 131

Abstract 131

Introduction 132

Experimental 133

Materials and Methods 133

Spectroscopic Measurements 133

v

Results and Discussion 134

W/Vis and CD Spectra 134

RR spectra 142

Conclusions 152

Acknowledgement 152

References 152

CHAPTER 8. STUDY ON THE STABILITY AND ELECTROCHEMICAL BEHAVIOR OF YEAST ISO-1-CYTOCHROME C BY ELECTROCHEMICAL AND SURFACE- ENHANCED RESONANCE RAMAN SCATTERING TECHNIQUES 157

Abstract 157

Introduction 158

Experimental 160

Materials 160

Apparatus and Methods 160

Results 162

Discussion 170

Conclusions 177

Acknowledgement 178

References 178

CHAPTER 9. GENEIL4L CONCLUSIONS 181

ACKNOWLEDGEMENTS 184

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1

Surface plasmon enhanced interracial electron transfer and resonance Raman,

surface-enhanced resonance Raman studies of cytochrome c mutants

Junwei Zheng

Major Professor: Edward S. Yeung

Surface plasmon resonance was utilized to enhance the electron transfer at

silver/solution interfaces. Photoelectrochemical reductions of nitrite, nitrate, and COZwere

studied on electrochemically roughened silver electrode surfaces. The dependence of the

photocument on photon energy, applied potential and concentration of nitrite demonstrates

that the photoelectrochemical reduction proceeds via photoemission process followed by the

capture of hydrated electrons. The excitation of plasmon resonances in nanosized metal

structures resulted in the enhancement of the photoemission process. In the case of

photoelectrocatalytic reduction of COZ,large photoelectrocatalytic effect for the reduction of

COZwas observed in the presence of surface adsorbed methylviologe~ which fi.mctionsas a

mediator for the photoexcited electron transfer from silver metal to C02 in solution.

Photoinduced reduction of microperoxidase-1 1 adsorbed on roughened silver electrode was

also observed and attributed to the direct photoejection of free electrons of silver metal.

Surface plasmon assisted electron transfer at nanostmctured silver particle surfaces was fbrther

determined by EPR method.

Resonance Raman studies of cytochrome c and its mutants demonstrate the sensitivity of

the spectra to mutations that affect the interactions of the heme peripheral substituents with the

protein matrix. The most dramatic differences in the spectra of the mutant cytochromes, as

compared with that of wild-type cytochrome c, occurred in the low-wavenumber region. The

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bands that are most strongly affected include bending modes of the thioether linkages and

propionic acid side-chains. Oxidation state dependent axial ligand switching of heme iron in

yeast iso-1-cytochrome c mutant F82H was determined by W-visible, circular dichroism and

resonance Raman spectra. From the analysis of the spectr~ it was concluded that in the

oxidized F82H axial Iigands to the heme iron are His-18 and Hk-82 whereas in the reduced

form the sixth ligand switches from His-82 to Met-80. The mutant possesses less distorted

porphyrin macrocycle and more opened conformation relative to that of wild type protein. The

comparisons of the sutiace-enhanced resonance Raman scattering spectra and cyclic

vokammetries of the proteins reveal that F82H mutant has more stable conformation and

negative redox reaction potential.

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1

CHAPTER 1

GENERAL INTRODUCTION

Surface plasmon of nanostructured metal particles has received increasing attention

and been extensively studied in recent years. A number of optical phenomen~ including

Surface-enhanced Raman Scattering (SERS)l-8, surface-enhanced absorption and

luminescence, second Harmonic generation (SHG)lO,can be strongly enhanced when

molecules adsorbed on roughened metal surfaces or on surfaces of nanostructured metal

particles. Those enhancements have been attributed to the excitation of the collective

electron oscillations or surface plasmon oscillations that engender huge electromagnetic field

both inside and out side of small metal particles. The fields inside result in strong absorption

of radiant energy. The fields outside, on other hand, stimulate enhanced optical emissions,

which are then fiuther enhanced by resonant interaction with particle at the shifted frequency.

Beyond the surface enhanced optical phenomen~ the enhancement due to surface plasmon

resonance has also recently been observed in photochemistry 1. One of the specific

objectives of this dissertation was foutiold on utilization of surface plasmon resonance to

enhance the electron transfer at silver/solution interfaces. The photoinduced electrochemical

reductions of nitrite, C02, microperoxidase-11 as a mo