“Switch I” mutant forms of the bacterial enhancer-binding protein NtrC that perturb the response to DNA

AUTOR(ES)

Yan, Dalai

FONTE

The National Academy of Sciences

RESUMO

NtrC (nitrogen regulatory protein C) is a bacterial enhancer-binding protein of 469 residues that activates transcription by σ54-holoenzyme. A region of its transcriptional activation (central) domain that is highly conserved among homologous activators of σ54-holoenzyme—residues 206–220—is essential for interaction with this RNA polymerase: it is required for contact with the polymerase and/or for coupling the energy from ATP hydrolysis to a change in the conformation of the polymerase that allows it to form transcriptionally productive open complexes. Several mutant NtrC proteins with amino acid substitutions in this region, including NtrCA216V and NtrCG219K, have normal ATPase activity but fail in transcriptional activation. We now report that other mutant forms carrying amino acid substitutions at these same positions, NtrCA216C and NtrCG219C, are capable of activating transcription when they are not bound to a DNA template (non-DNA-binding derivatives with an altered helix–turn–helix DNA-binding motif at the C terminus of the protein) but are unable to do so when they are bound to a DNA template, whether or not it carries a specific enhancer. Enhancer DNA remains a positive allosteric effector of ATP hydrolysis, as it is for wild-type NtrC but, surprisingly, appears to have become a negative allosteric effector for some aspect of interaction with σ54-holoenzyme. The conserved region in which these amino acid substitutions occur (206–220) is equivalent to the Switch I region of a large group of purine nucleotide-binding proteins. Interesting analogies can be drawn between the Switch I region of NtrC and that of p21ras.

Documentos Relacionados

• MgATP Binding and Hydrolysis Determinants of NtrC, a Bacterial Enhancer-Binding Protein
• Physical evidence for a phosphorylation-dependent conformational change in the enhancer-binding protein NtrC
• Active contribution of two domains to cooperative DNA binding of the enhancer-binding protein nitrogen regulator I (NtrC) of Escherichia coli: stimulation by phosphorylation and the binding of ATP.
• Repressor Mutant Forms of the Azospirillum brasilense NtrC Protein
• Purification of the c-fos enhancer-binding protein.