Engagement of Arginine Finger to ATP Triggers Large Conformational Changes in NtrC1 AAA+ ATPase for Remodeling Bacterial RNA Polymerase

Abstract

SUMMARY The NtrC-like AAA+ ATPases control virulence and other important bacterial activities through delivering mechanical work to σ54-RNA polymerase to activate transcription from σ54-dependent genes. We report the first crystal structure for such an ATPase, NtrC1 of Aquifex aeolicus, in which the catalytic arginine engages the gamma-phosphate of ATP. Comparing the new structure with those previously known for apo and ADP-bound states supports a novel rigid-body displacement model that is consistent with large-scale conformational changes observed by low-resolution methods. First, the arginine-finger induces rigid-body roll, extending surface loops above the plane of the ATPase ring to bind σ54. Second, ATP hydrolysis permits Pi release and retraction of the arginine with a reversed roll, remodeling σ54-RNAP. This model provides a fresh perspective on how ATPase subunits interact within the ring-ensemble to promote transcription, directing attention to structural changes on the arginine-finger side of an ATP-bound interface.

Nucleotide-induced asymmetry within ATPase activator ring drives σ54–RNAP interaction and ATP hydrolysis

“…5). Although it was observed earlier that s54 protein can bind with other oligomeric states of the bEBPs (Rappas et al 2005;Chen et al 2010), we propose that the asymmetric hexamer provides a unique configuration for guiding the protein-protein interaction. Our preliminary EM data for the NtrC1 ATPase in complex with s54-promoter-RNAP suggest the presence of a similarly asymmetric NtrC1 hexamer (S Chowdhury, S DeCarlo, and BT Nixon, unpubl.).…”

“…What does appear certain is that all of the multimeric ATPases need to break rotational symmetry or lower it in order to function efficiently. That requirement may explain inhibition of the ATPase activity by saturating amounts of ATP in the bEBPs PspF and NtrC1 (Schumacher et al 2008;Chen et al 2010).…”

“…Several bEBPs have been crystallized as hexamers and heptamers, but no mechanism for hydrolysis has yet emerged from those structures (Lee et al 2003;Rappas et al 2005Rappas et al , 2006Sallai and Tucker 2005;Batchelor et al 2008;Chen et al 2010). We showed previously that binding of ATP but not ADP causes large structural changes in the bEBP NtrC1 from the extreme hyperthermophile Aquifex aeolicus (Chen et al 2007.…”

Spontaneous Reversions of an Evolutionary Trait Loss Reveal Regulators of a Small RNA That Controls Multicellular Development in Myxobacteria

“…The altered amino acid is located next to an annotated arginine finger (Arg311) and within an annotated 54 interaction domain. Binding of the arginine finger to ATP triggers a large conformational shift, which is essential for interaction with 54 -RNA polymerase (40). Thus, the mutation of Tyr310 may negatively affect the interaction with 54 -RNA polymerase, resulting in the absence of pxr transcription.…”

IBMPFD and p97, the Structural and Molecular Basis for Functional Disruption

“…Such methods include cryo EM and SAXS. Cryo EM has revealed flexibility in the N-domains of E. coli AAA + protein ClpA (Ishikawa et al, 2004) and SAXS experiments have shown large conformational changes in NtrC1 (Chen et al, 2010). Although in some cases conformational changes observed with different methods do not completely agree, it is widely accepted that AAA + proteins undergo dynamic movements during their catalytic cycle.…”