Nuclear Magnetic Resonance Structure and Dynamics of the Response Regulator Sma0114 from Sinorhizobium meliloti

Abstract

Receiver domains control intracellular responses triggered by signal transduction in bacterial two-component systems. Here, we report the solution NMR structure and dynamics of Sma0114 from the bacterium Sinorhizobium meliloti, the first such characterization of a receiver domain from the HWE-kinase family of two-component systems. The structure of Sma0114 adopts a prototypical α5/β5 Rossman-fold but has features that set it apart from other receiver domains. The fourth β-strand of Sma0114 houses a PFxFATGY sequence motif, common to many HWE-kinase-associated receiver domains. This sequence motif in Sma0114 may substitute for the conserved Y-T coupling mechanism, which propagates conformational transitions in the 455 (α4-β5-α5) faces of receiver domains, to prime them for binding downstream effectors once they become activated by phosphorylation. In addition, Sma0114 lacks the fourth α-helix of the consensus 455 face and 15N relaxation data show that it is replaced by a segment that is flexible on the ps-ns timescale. Secondary structure prediction of Sma0114 and other HWE-kinase-associated receiver domains suggests that the absence of helix α4 may be a conserved property of this family. In spite of these differences, Sma0114 has a conserved active site, binds divalent metal ions such as Mg2+ and Ca2+ that are required for phosphorylation, and exhibits μs-ms active site dynamics similar to other receiver domains. Taken together, our results suggest that Sma0114 has a conserved active site but differs from typical receiver domains in the structure of the 455 face that is used to effect signal transduction following activation.

Role of the PFXFATG[G/Y] Motif in the Activation of SdrG, a Response Regulator Involved in the Alphaproteobacterial General Stress Response

“…The 20 structures of the NMR ensemble superimposed with a backbone atoms root-mean-square deviation (rmsd) of 0.51 Å (Figures 3A and 3B). Compared with the Rossman fold of receiver domains, the SdrG structure revealed a divergent fold where the helix a4 (the nomenclature of secondary structures of regular REC has been conserved throughout the manuscript) is replaced by a flexible segment as was already observed in the structure of Sma0114 (Sheftic et al, 2012). This probably represents a hallmark of this response regulator family.…”

“…Instead, SdrG shows a derivation of a conserved PFXFATGY motif (PFXFATGG) that includes the threonine implicated in Y-T coupling in prototypical response regulators. Response regulators with such a motif, also called ''FAT GUY'' response regulators, were only recently described (Sheftic et al, 2011(Sheftic et al, , 2012 and are often associated with the HWE family of histidine kinases (Karniol and Vierstra, 2004;Sheftic et al, 2014), to which some of the Paks belong. The structures of one FAT GUY response regulator, Sma0114 of Sinorhizobium meliloti, were solved (Sheftic et al, 2011(Sheftic et al, , 2012(Sheftic et al, , 2014, but the role of the conserved PFXFATGY motif was not determined in these studies.…”

Colocalization of Fast and Slow Timescale Dynamics in the Allosteric Signaling Protein CheY

“…3, 22-24 While fast backbone dynamics have also been reported, they have not been shown to be involved in the allosteric transition. 3, 24, 25 Here, we report the first multi-timescale study of both an unphosphorylated and phosphorylated RR including measurement of fast side-chain dynamics.…”

A single-domain response regulator functions as integrating hub to coordinate general stress response and development in alpha-proteobacteria

“…In contrast to prototypical response regulators, MrrA lacks one of the residues involved in the Y-T coupling mechanism required for intramolecular signal transduction of Rec domains. Instead, it harbors the recently described FATGUY motif (35,51,52). Three other FATGUY response regulators are described, SdrG of S. melonis, Mext_0407 of Methylobacterium extorquens and Sma0114 of Sinorhizobium meliloti (34,53,54).…”