Mechanism of induction of the homologous global carbon regulators YvoA and DasR in bacteria
Final Report Abstract
Members of the GntR/HutC family of bacterial repressors constitute so-called one-component signal transduction elements and in case of DasR from S. coelicolor and NagR from B. subtilis link environmental stimuli to cellular responses. Although the GntR proteins represent a large family of repressors, only sparse knowledge was available on the atomic mechanisms that regulate their function. In order to obtain an atomic understanding, insight has not only to be gained into the determinants for specific DNA and effector recognition but also into the allosteric coupling mechanism that allows for the modulation of the DNA-binding affinities of the repressors upon binding of small molecule effectors with the sterically distant effector-binding domains (EBDs). The central aims for the second funding period of the project were: I: How do these repressors accomplish the highly specific recognition of DNA operator sites? II: What are the molecular determinants for the specific recognition of small effector molecules? III: What is the nature of the allosteric switch through which binding of the effector molecule to the remote effector-binding site abrogates DNA binding in the N-terminal DNA-binding domains (DBDs)? Insight into aim I and II was obtained from a total of 8 crystal structure determinations and additional biochemical and computational studies that revealed the atomic determinants of specific dre-site DNA recognition and also of the specific interaction with the effectors N-acetylglucosamine-6-phosphate (GlcNAc-6-P) and glucosamine-6-phosphate (GlcN-6-P). Together with molecular dynamics simulations and a comparison with other GntR/HutC family members these data allowed us to propose a detailed model for the allosteric regulation of these repressors (aim III). This behavior appears best described by a conformational selection model. In effector-free DasR and NagR a heightened flexibility in the EBDs enables the DBDs to sample a huge variety of different orientations (state I) and among these also a DNA-binding competent conformation (state II). Effector binding to the EBDs of DasR significantly reorganizes the atomic structure of the latter. However, rather than locking the DBDs into a specific orientation, effector-induced formation of β-strand β* in the DBD-EBD-linker segment merely appears to take the DBDs ‘on a shorter leash’ (state III) thereby impeding the ‘downwards’ positioning of the DBDs that is necessary for the concerted binding of the two DBDs to operator DNA (state V versus state II).
Publications
- (2015) Multiple allosteric effectors control the affinity of DasR for its target sites. Biochem Biophys Res Commun. 464: 324-329
Tenconi E., Urem M., Świątek-Połatyńska M. A., Titgemeyer F., Muller Y. A., van Wezel G. P. & Rigali S.
(See online at https://doi.org/10.1016/j.bbrc.2015.06.152) - (2015). Structural insight into operator dre-sites recognition and effector binding in the GntR/HutC transcription regulator NagR. Nucleic Acids Research 43: 1283-1296
Fillenberg, S. B., Grau, F. C., Seidel, G. & Muller, Y. A.
(See online at https://doi.org/10.1093/nar/gku1374) - Crystal structures of the global regulator DasR from Streptomyces coelicolor: Implications for the allosteric regulation of GntR/HutC repressors. PLoS ONE 11(6): e0157691, 2016
Fillenberg, S. B., Friess, M., Körner, S., Böckmann, R. A. & Muller, Y. A.
(See online at https://doi.org/10.1371/journal.pone.0157691)
