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Probing the orientation of electrostatically immobilized cytochrome C by time of flight secondary ion mass spectrometry and sum frequency generation spectroscopy
By taking advantage of the electron pathway through the heme group in cytochrome c (CytoC) electrochemists have built sensors based upon CytoC immobilized onto metal electrodes. Previous studies have shown that the electron transfer rate through the protein is a function of the position of this heme...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Vacuum Society
2013
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4000547/ https://www.ncbi.nlm.nih.gov/pubmed/24706131 http://dx.doi.org/10.1186/1559-4106-8-18 |
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author | Baio, Joe E Weidner, Tobias Ramey, Dennis Pruzinsky, Leah Castner, David G |
author_facet | Baio, Joe E Weidner, Tobias Ramey, Dennis Pruzinsky, Leah Castner, David G |
author_sort | Baio, Joe E |
collection | PubMed |
description | By taking advantage of the electron pathway through the heme group in cytochrome c (CytoC) electrochemists have built sensors based upon CytoC immobilized onto metal electrodes. Previous studies have shown that the electron transfer rate through the protein is a function of the position of this heme group with respect to the electrode surface. In this study a detailed examination of CytoC orientation when electrostatically immobilized onto both amine (NH(3)(+)) and carboxyl (COO(-)) functionalized gold is presented. Protein coverage, on both surfaces, was monitored by the change in the atomic % N, as determined by x-ray photoelectron spectroscopy. Spectral features within the in situ sum frequency generation vibrational spectra, acquired for the protein interacting with positively and negatively charged surfaces, indicates that these electrostatic interactions do induce the protein into a well ordered film. Time of flight secondary ion mass spectrometry data demonstrated a clear separation between the two samples based on the intensity differences of secondary ions stemming from amino acids located asymmetrically within CytoC (cysteine: C(2)H(6)NS(+); glutamic acid: C(4)H(6)NO(+) and C(4)H(8)NO(2)(+); leucine: C(5)H(12)N(+)). For a more quantitative examination of orientation, we developed a ratio comparing the sum of the intensities of secondary-ions stemming from the amino acid residues at either end of the protein. The 50 % increase in this ratio, observed between the protein covered NH(3)(+) and COO(-) substrates, indicates opposite orientations of the CytoC on the two different surfaces. |
format | Online Article Text |
id | pubmed-4000547 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2013 |
publisher | American Vacuum Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-40005472014-12-01 Probing the orientation of electrostatically immobilized cytochrome C by time of flight secondary ion mass spectrometry and sum frequency generation spectroscopy Baio, Joe E Weidner, Tobias Ramey, Dennis Pruzinsky, Leah Castner, David G Biointerphases Article By taking advantage of the electron pathway through the heme group in cytochrome c (CytoC) electrochemists have built sensors based upon CytoC immobilized onto metal electrodes. Previous studies have shown that the electron transfer rate through the protein is a function of the position of this heme group with respect to the electrode surface. In this study a detailed examination of CytoC orientation when electrostatically immobilized onto both amine (NH(3)(+)) and carboxyl (COO(-)) functionalized gold is presented. Protein coverage, on both surfaces, was monitored by the change in the atomic % N, as determined by x-ray photoelectron spectroscopy. Spectral features within the in situ sum frequency generation vibrational spectra, acquired for the protein interacting with positively and negatively charged surfaces, indicates that these electrostatic interactions do induce the protein into a well ordered film. Time of flight secondary ion mass spectrometry data demonstrated a clear separation between the two samples based on the intensity differences of secondary ions stemming from amino acids located asymmetrically within CytoC (cysteine: C(2)H(6)NS(+); glutamic acid: C(4)H(6)NO(+) and C(4)H(8)NO(2)(+); leucine: C(5)H(12)N(+)). For a more quantitative examination of orientation, we developed a ratio comparing the sum of the intensities of secondary-ions stemming from the amino acid residues at either end of the protein. The 50 % increase in this ratio, observed between the protein covered NH(3)(+) and COO(-) substrates, indicates opposite orientations of the CytoC on the two different surfaces. American Vacuum Society 2013-12 2013-07-26 /pmc/articles/PMC4000547/ /pubmed/24706131 http://dx.doi.org/10.1186/1559-4106-8-18 Text en © 2013 Baio et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Article Baio, Joe E Weidner, Tobias Ramey, Dennis Pruzinsky, Leah Castner, David G Probing the orientation of electrostatically immobilized cytochrome C by time of flight secondary ion mass spectrometry and sum frequency generation spectroscopy |
title | Probing the orientation of electrostatically immobilized cytochrome C by time of flight secondary ion mass spectrometry and sum frequency generation spectroscopy |
title_full | Probing the orientation of electrostatically immobilized cytochrome C by time of flight secondary ion mass spectrometry and sum frequency generation spectroscopy |
title_fullStr | Probing the orientation of electrostatically immobilized cytochrome C by time of flight secondary ion mass spectrometry and sum frequency generation spectroscopy |
title_full_unstemmed | Probing the orientation of electrostatically immobilized cytochrome C by time of flight secondary ion mass spectrometry and sum frequency generation spectroscopy |
title_short | Probing the orientation of electrostatically immobilized cytochrome C by time of flight secondary ion mass spectrometry and sum frequency generation spectroscopy |
title_sort | probing the orientation of electrostatically immobilized cytochrome c by time of flight secondary ion mass spectrometry and sum frequency generation spectroscopy |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4000547/ https://www.ncbi.nlm.nih.gov/pubmed/24706131 http://dx.doi.org/10.1186/1559-4106-8-18 |
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