Oligomer formation of the bacterial second messenger c-di-GMP : reaction rates and equilibrium constants indicate a monomeric state at physiological concentrations

Gentner, Martin and Allan, Martin G. and Zaehringer, Franziska and Schirmer, Tilman and Grzesiek, Stephan. (2012) Oligomer formation of the bacterial second messenger c-di-GMP : reaction rates and equilibrium constants indicate a monomeric state at physiological concentrations. The journal of the American Chemical Society, 134 (2). pp. 1019-1029.

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Cyclic di-guanosine-monophosphate (c-di-GMP) is a bacterial signaling molecule that triggers a switch from motile to sessile bacterial lifestyles. This mechanism is of considerable pharmaceutical interest, since it is related to bacterial virulence, biofilm formation and persistence of infection. Previously, c-di-GMP has been reported to display a rich polymorphism of various oligomeric forms at millimolar concentrations, which differ in base stacking and G-quartet interactions. Here, we have analyzed the equilibrium and exchange kinetics between these various forms by NMR spectroscopy. We find that the association of the monomer into a dimeric form is in fast exchange (> milliseconds) with an equilibrium constant of about 1 mM. At concentrations above 100 ?M higher oligomers are formed in the presence of cations. These are presumably tetramers and octamers, with octamers dominating above about 0.5 mM. Thus at the low micromolar concentrations of the cellular environment and in the absence of additional compounds that stabilize oligomers, c-di-GMP should be predominantly monomeric. This finding has important implications for the understanding of c-di-GMP recognition by protein receptors. In contrast to the monomer/dimer exchange, formation and dissociation of higher oligomers occurs on a timescale of several hours to days. The time course can be described quantitatively by a simple kinetic model where tetramers are intermediates of octamer formation. The extremely slow oligomer dissociation may generate severe artifacts in biological experiments when c-di-GMP is diluted from concentrated stock solution. We present a simple method to quantify c-di-GMP monomers and oligomers from UV spectra and a procedure to dissolve the unwanted oligomers by an annealing step.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Former Organization Units Biozentrum > Structural Biology (Schirmer)
UniBasel Contributors:Grzesiek, Stephan and Schirmer, Tilman and Gentner, Martin and Zähringer, Franziska Ursula
Item Type:Article, refereed
Article Subtype:Research Article
Publisher:American Chemical Society
Note:Publication type according to Uni Basel Research Database: Journal article
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Last Modified:31 Dec 2015 10:50
Deposited On:11 Oct 2012 15:14

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