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J Biol Chem. 2015 Jan 30;290(5):3069-80. doi: 10.1074/jbc.M114.619619. Epub 2014 Nov 28.

Identification, characterization, and structure analysis of the cyclic di-AMP-binding PII-like signal transduction protein DarA.

Author information

1
From the Departments of General Microbiology.
2
Molecular Structural Biology, and.
3
Molecular Enzymology, Georg August University Göttingen, 37077 Göttingen, Germany.
4
Department of Functional Genomics, Interfaculty Institute for Genetics and Functional Genomics, University Medicine Greifswald, 17487 Greifswald, Germany.
5
BIOLOG Life Science Institute, 28199 Bremen, Germany, and.
6
Research Core Unit Metabolomics, Hannover Medical School, 30625 Hannover, Germany.
7
From the Departments of General Microbiology, jstuelk@gwdg.de.
8
Molecular Structural Biology, and rficner@uni-goettingen.de.

Abstract

The cyclic dimeric AMP nucleotide c-di-AMP is an essential second messenger in Bacillus subtilis. We have identified the protein DarA as one of the prominent c-di-AMP receptors in B. subtilis. Crystal structure analysis shows that DarA is highly homologous to PII signal transducer proteins. In contrast to PII proteins, the functionally important B- and T-loops are swapped with respect to their size. DarA is a homotrimer that binds three molecules of c-di-AMP, each in a pocket located between two subunits. We demonstrate that DarA is capable to bind c-di-AMP and with lower affinity cyclic GMP-AMP (3'3'-cGAMP) but not c-di-GMP or 2'3'-cGAMP. Consistently the crystal structure shows that within the ligand-binding pocket only one adenine is highly specifically recognized, whereas the pocket for the other adenine appears to be promiscuous. Comparison with a homologous ligand-free DarA structure reveals that c-di-AMP binding is accompanied by conformational changes of both the fold and the position of the B-loop in DarA.

KEYWORDS:

Bacterial Signal Transduction; Crystal Structure; Cyclic Di-GMP (c-di-GMP); Cyclic Diadenosine Monophosphate (c-di-AMP); Isothermal Titration Calorimetry (ITC)

PMID:
25433025
PMCID:
PMC4317042
DOI:
10.1074/jbc.M114.619619
[Indexed for MEDLINE]
Free PMC Article

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