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Analysis of c-di-GMP Levels Synthesized by a Photoreceptor Protein in Response to Different Light Qualities Using an In Vitro Enzymatic Assay

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c-di-GMP Signaling

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1657))

Abstract

Diguanylate cyclases are enzymes that use two GTP molecules to produce one molecule cyclic dimeric guanosine monophosphate (c-di-GMP). This cyclic dinucleotide is an ubiquitous prokaryotic second messenger that controls a variety of cell functions. Several proteins have been described which contain a photoreceptor domain fused to a diguanylate cyclase. The cyanobacterial light sensor Cph2 is responsible for the blue-light induced synthesis of c-di-GMP in Synechocystis sp. PCC 6803. Here, we provide a detailed protocol for an in vitro enzymatic assay with a purified photoreceptor protein using light as the crucial reaction parameter for c-di-GMP synthesis. The assay is accomplished under continuous illumination with light of different quality with inactivation of the enzyme by heat denaturation. Analytics are performed using HPLC-UV.

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Acknowledgment

V.A. acknowledges funding by the Jürgen-Manchot-Stiftung.

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Authors and Affiliations

  1. Institute of Biology III, Albert-Ludwigs-University Freiburg, Schänzlestr. 1, 79104, Freiburg, Germany

    Veronika Angerer & Annegret Wilde

  2. Department of Chemistry, Philipps-University Marburg, 35032, Marburg, Germany

    Lars-Oliver Essen

Authors
  1. Veronika Angerer
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  2. Lars-Oliver Essen
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  3. Annegret Wilde
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Corresponding author

Correspondence to Annegret Wilde .

Editor information

Editors and Affiliations

  1. Department of Biological Sciences, Binghamton University Department of Biological Sciences, Binghamton, New York, USA

    Karin Sauer

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Angerer, V., Essen, LO., Wilde, A. (2017). Analysis of c-di-GMP Levels Synthesized by a Photoreceptor Protein in Response to Different Light Qualities Using an In Vitro Enzymatic Assay. In: Sauer, K. (eds) c-di-GMP Signaling. Methods in Molecular Biology, vol 1657. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7240-1_15

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  • DOI: https://doi.org/10.1007/978-1-4939-7240-1_15

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7239-5

  • Online ISBN: 978-1-4939-7240-1

  • eBook Packages: Springer Protocols

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