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SDS-PAGE and Dot Blot Autoradiography: Tools for Quantifying Histidine Kinase Autophosphorylation

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Histidine Phosphorylation

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2077))

Abstract

Histidine kinases play a vital role in bacterial signal transduction. However, methods for studying the activity of histidine kinases in vitro are limited in comparison to those for investigating serine, threonine, and tyrosine kinases, largely due to the lability of the phosphoramidate (P-N) bond. Here, we describe two useful methods for quantifying histidine kinase autophosphorylation: SDS-PAGE autoradiography and dot blot autoradiography/scintillation counting.

Jonathan T. Fischer and Ilana Heckler contributed equally to this work.

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Acknowledgments

This work was supported by NIH GM118894 and NSF CHE-1607532.

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Authors and Affiliations

  1. Department of Chemistry, Stony Brook University, Stony Brook, NY, USA

    Jonathan T. Fischer, Ilana Heckler & Elizabeth M. Boon

  2. Institute of Chemical Biology and Drug Discovery, Stony Brook, NY, USA

    Elizabeth M. Boon

Authors
  1. Jonathan T. Fischer
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  2. Ilana Heckler
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  3. Elizabeth M. Boon
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Corresponding author

Correspondence to Elizabeth M. Boon .

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Editors and Affiliations

  1. Department of Biochemistry, Centre for Proteome Research Institute of Integrative Biology, University of Liverpool, Liverpool, UK

    Claire E. Eyers

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Fischer, J.T., Heckler, I., Boon, E.M. (2020). SDS-PAGE and Dot Blot Autoradiography: Tools for Quantifying Histidine Kinase Autophosphorylation. In: Eyers, C. (eds) Histidine Phosphorylation. Methods in Molecular Biology, vol 2077. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9884-5_3

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  • DOI: https://doi.org/10.1007/978-1-4939-9884-5_3

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9883-8

  • Online ISBN: 978-1-4939-9884-5

  • eBook Packages: Springer Protocols

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