Abstract
LysR Type Transcriptional Regulators (LTTRs) regulate basic metabolic pathways or virulence gene expression in prokaryotes. Evidence suggests that the activation of LTTRs involves a conformational change from an inactive compact apo- configuration that represses transcription to an active, expanded holo- form that promotes it. However, no LTTR has yet been observed to adopt both configurations. Here, we report the results of structural studies of various forms of the LTTR DntR. Crystal structures of apo-DntR and of a partially autoinducing mutant H169T-DntR suggest that active and inactive DntR maintain a compact homotetrameric configuration. However, Small Angle X-ray Scattering (SAXS) studies on solutions of apo-, H169T- and inducer-bound holo-DntR indicate a different behaviour, suggesting that while apo-DntR maintains a compact configuration in solution both H169T- and holo-DntR adopt an expanded conformation. Models of the SAXS-obtained solution conformations of apo- and holo-DntR homotetramers in complex with promoter-operator region DNA are consistent with previous observations of a shifting of LTTR DNA binding sites upon activation and a consequent relaxation in the bend of the promoter-operator region DNA. Our results thus provide clear evidence at the molecular level which strongly supports the ‘sliding dimer’ hypothesis concerning LTTR activation mechanisms.
Highlights
LysR Type Transcriptional Regulators (LTTRs), the largest family of transcription factors found in prokaryotes[1,2], are involved in the regulation of basic metabolic pathways or virulence gene expression[3] and share a high degree of structural identity[4]
The current consensus is that LTTRs regulate transcription through large conformational changes which modify LTTR DNA binding sites promoter region DNA
The results presented here provide the first clear evidence both that a single LTTR homotetramer can adopt both conformations and that the conformation adopted by the homotetramer is a function of activation state
Summary
LysR Type Transcriptional Regulators (LTTRs), the largest family of transcription factors found in prokaryotes[1,2], are involved in the regulation of basic metabolic pathways or virulence gene expression[3] and share a high degree of structural identity[4]. The current hypothesis[6,7] is that upon activation LTTRs morph from a compact to an expanded homotetrameric configuration (Fig. 1b–d)) allowing the DBD dimers to bind to different DNA sites. Putative models bound to promoter DNA regions appear to support the physiological relevance of the solution conformations of apo- and holo-DntR obtained. These models suggest that a shift of binding sites, accompanied by a relaxation of DNA bend, would occur upon activation of DntR. Our results show that a single LTTR can adopt different conformations, depending on activation state, providing compelling structural evidence to support the sliding dimer mechanism for the activation of homotetrameric LTTRs and confirming that this involves a change in quaternary structure from compact to expanded conformations
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