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A “How-To” Guide to the Stark Spectroscopy of Flavins and Flavoproteins

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Flavins and Flavoproteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1146))

Abstract

Flavins and flavoproteins have been studied by a plethora of spectroscopic techniques. Beginning with the characterization of DNA photolyases and the discovery of the diversity of roles played by excited-state flavins in photobiology, the characterization of the electronic excited state of flavins has become increasingly important. In this protocol, we provide a guide to using Stark spectroscopy in obtaining the degree of electronic charge redistribution in simple flavins and in flavoproteins. Stark spectroscopy is technically simpler than more common approaches used to explore the structure of the excited state, considerably cheaper to implement, and yet very powerful in its scope. At the end of this guide, we present data taken on non-photobiological flavoproteins, glutathione reductase and lipoamide dehydrogenase, that suggest that Stark spectroscopy is a unique way to elucidate the electrostatic environment that the flavin cofactor experiences bound inside the protein.

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Acknowledgements

Dr. Goutham Kodali gave very useful feedback on this manuscript. We wish to thank Dr. Ron Koder for the TPARF and Dr. Nancy Hopkins for the lipoamide dehydrogenase enzyme. R.P. and R.J.S. were supported by a grant from the NSF Division of Chemistry (CHE-0847855). R.J.S. is grateful for support from the NSF Molecular Biosciences Division (MCB-0347087).

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Authors and Affiliations

  1. Department of Chemistry, Temple University, 250B Beury Hall, 1901N. 13th Street, Philadelphia, PA, 19122, USA

    Raymond F. Pauszek & Robert J. Stanley

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  1. Raymond F. Pauszek
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  2. Robert J. Stanley
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Correspondence to Robert J. Stanley .

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Editors and Affiliations

  1. Institut für Physikalische Chemie, Albert-Ludwigs-Universität Freiburg, Germany

    Stefan Weber

  2. Institut für Physikalische Chemie, Albert-Ludwigs-Universität Freiburg, Germany

    Erik Schleicher

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Pauszek, R.F., Stanley, R.J. (2014). A “How-To” Guide to the Stark Spectroscopy of Flavins and Flavoproteins. In: Weber, S., Schleicher, E. (eds) Flavins and Flavoproteins. Methods in Molecular Biology, vol 1146. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0452-5_17

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  • DOI: https://doi.org/10.1007/978-1-4939-0452-5_17

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-0451-8

  • Online ISBN: 978-1-4939-0452-5

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