Chapter title |
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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Chapter number | 15 |
Book title |
c-di-GMP Signaling
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Published in |
Methods in molecular biology, January 2017
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DOI | 10.1007/978-1-4939-7240-1_15 |
Pubmed ID | |
Book ISBNs |
978-1-4939-7239-5, 978-1-4939-7240-1
|
Authors |
Veronika Angerer, Lars-Oliver Essen, Annegret Wilde |
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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