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Capture compound mass spectrometry - a powerful tool to identify novel c-di-GMP effector proteins

Laventie, Benoît-Joseph and Nesper, Jutta and Ahrné, Erik and Glatter, Timo and Schmidt, Alexander and Jenal, Urs. (2015) Capture compound mass spectrometry - a powerful tool to identify novel c-di-GMP effector proteins. Journal of visualized experiments, H. 97 , e51404.

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Official URL: http://edoc.unibas.ch/dok/A6438751

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Abstract

Considerable progress has been made during the last decade towards the identification and characterization of enzymes involved in the synthesis (diguanylate cyclases) and degradation (phosphodiesterases) of the second messenger c-di-GMP. In contrast, little information is available regarding the molecular mechanisms and cellular components through which this signaling molecule regulates a diverse range of cellular processes. Most of the known effector proteins belong to the PilZ family or are degenerated diguanylate cyclases or phosphodiesterases that have given up on catalysis and have adopted effector function. Thus, to better define the cellular c-di-GMP network in a wide range of bacteria experimental methods are required to identify and validate novel effectors for which reliable in silico predictions fail. We have recently developed a novel Capture Compound Mass Spectrometry (CCMS) based technology as a powerful tool to biochemically identify and characterize c-di-GMP binding proteins. This technique has previously been reported to be applicable to a wide range of organisms(1). Here we give a detailed description of the protocol that we utilize to probe such signaling components. As an example, we use Pseudomonas aeruginosa, an opportunistic pathogen in which c-di-GMP plays a critical role in virulence and biofilm control. CCMS identified 74% (38/51) of the known or predicted components of the c-di-GMP network. This study explains the CCMS procedure in detail, and establishes it as a powerful and versatile tool to identify novel components involved in small molecule signaling.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Growth & Development
05 Faculty of Science > Departement Biozentrum > Infection Biology > Molecular Microbiology (Jenal)
05 Faculty of Science > Departement Biozentrum > Growth & Development > Molecular Microbiology (Jenal)
05 Faculty of Science > Departement Biozentrum > Infection Biology
UniBasel Contributors:Jenal, Urs and Laventie, Benoit-Joseph and Nesper, Jutta and Ahrné, Erik and Schmidt, Alexander
Item Type:Article, refereed
Article Subtype:Research Article
Bibsysno:Link to catalogue
Publisher:JoVE
ISSN:1940-087X
Note:Publication type according to Uni Basel Research Database: Journal article
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Last Modified:06 Nov 2015 10:21
Deposited On:06 Nov 2015 10:21

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