Probing the activity of diguanylate cyclases and c-di-GMP phosphodiesterases in real-time by CD spectroscopy. - Archive ouverte HAL Access content directly
Journal Articles Nucleic Acids Research Year : 2013

Probing the activity of diguanylate cyclases and c-di-GMP phosphodiesterases in real-time by CD spectroscopy.

(1) , (1) , (1) , (1) , (1) , (2) , (1) , (2, 3) , (1)
1
2
3

Abstract

Bacteria react to adverse environmental stimuli by clustering into organized communities called biofilms. A remarkably sophisticated control system based on the dinucleotide 3'-5' cyclic diguanylic acid (c-di-GMP) is involved in deciding whether to form or abandon biofilms. The ability of c-di-GMP to form self-intercalated dimers is also thought to play a role in this complex regulation. A great advantage in the quest of elucidating the catalytic properties of the enzymes involved in c-di-GMP turnover (diguanylate cyclases and phosphodiesterases) would come from the availability of an experimental approach for in vitro quantification of c-di-GMP in real-time. Here, we show that c-di-GMP can be detected and quantified by circular dichroism (CD) spectroscopy in the low micromolar range. The method is based on the selective ability of manganese ions to induce formation of the intercalated dimer of the c-di-GMP dinucleotide in solution, which displays an intense sigmoidal CD spectrum in the near-ultraviolet region. This characteristic spectrum originates from the stacking interaction of the four mutually intercalated guanines, as it is absent in the other cyclic dinucleotide 3'-5' cyclic adenilic acid (c-di-AMP). Thus, near-ultraviolet CD can be used to effectively quantify in real-time the activity of diguanylate cyclases and phosphodiesterases in solution.
Fichier principal
Vignette du fichier
Stelitano_Nucl_A_Res.pdf (3.53 Mo) Télécharger le fichier
Origin : Publication funded by an institution
Loading...

Dates and versions

pasteur-01048743 , version 1 (25-07-2014)

Identifiers

Cite

Valentina Stelitano, Annegret Brandt, Silvia Fernicola, Stefano Franceschini, Giorgio Giardina, et al.. Probing the activity of diguanylate cyclases and c-di-GMP phosphodiesterases in real-time by CD spectroscopy.. Nucleic Acids Research, 2013, 41 (7), pp.e79. ⟨10.1093/nar/gkt028⟩. ⟨pasteur-01048743⟩

Collections

RIIP RIIP_FCB
23 View
156 Download

Altmetric

Share

Gmail Facebook Twitter LinkedIn More