PMID:20582742
| Citation |
Zähringer, F, Massa, C and Schirmer, T (2011) Efficient enzymatic production of the bacterial second messenger c-di-GMP by the diguanylate cyclase YdeH from E. coli. Appl. Biochem. Biotechnol. 163:71-9 |
|---|---|
| Abstract |
Cyclic di-GMP (c-di-GMP) is an almost universal bacterial second messenger involved in the regulation of cell surface-associated traits and the persistence of infections. GGDEF and EAL domain-containing proteins catalyse c-di-GMP synthesis and degradation, respectively. We report the enzymatic large-scale synthesis of c-di-GMP, making use of the GGDEF domain-containing protein YdeH from Escherichia coli. Overexpression and purification of YdeH have been established, and the conditions for c-di-GMP synthesis were optimised. In contrast to the chemical synthesis of c-di-GMP, enzymatic c-di-GMP production is a one-step reaction that can easily be performed with the equipment of a standard biochemical lab. The protocol allows the production of milligram amounts of c-di-GMP within 1 day and paves the way for extensive biochemical and biophysical studies on c-di-GMP-mediated processes. |
| Links |
PubMed Online version:10.1007/s12010-010-9017-x |
| Keywords |
Cyclic GMP/biosynthesis; Escherichia coli/enzymology; Escherichia coli/genetics; Escherichia coli/metabolism; Escherichia coli Proteins/chemistry; Escherichia coli Proteins/genetics; Escherichia coli Proteins/isolation & purification; Escherichia coli Proteins/metabolism; Gene Expression Regulation, Bacterial; Phosphorus-Oxygen Lyases/chemistry; Phosphorus-Oxygen Lyases/genetics; Phosphorus-Oxygen Lyases/isolation & purification; Phosphorus-Oxygen Lyases/metabolism; Protein Structure, Tertiary; Second Messenger Systems |
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