Large-scale filament formation inhibits the activity of CTP synthetase.
Bottom Line: Structure-guided mutagenesis and mathematical modeling further indicate that coupling activity to polymerization promotes cooperative catalytic regulation.This previously uncharacterized regulatory mechanism is important for cellular function since a mutant that disrupts CtpS polymerization disrupts E. coli growth and metabolic regulation without reducing CTP levels.We propose that regulation by large-scale polymerization enables ultrasensitive control of enzymatic activity while storing an enzyme subpopulation in a conformationally restricted form that is readily activatable.
Affiliation: Department of Molecular Biology, Princeton University, Princeton, United States.Show MeSH
Mentions: The cryo-EM structure of the CtpS filament offers insight into the mechanism of enzymatic regulation. All of the enzyme active sites are solvent accessible, suggesting that UTP, ATP, and glutamine can freely diffuse into the filament (Figure 5A). This observation rules out occlusion of active sites as a regulatory mechanism. An alternative mechanism of CtpS inhibition is blocking the transfer of ammonia between the GATase and ALase active sites, which are separated by ∼25 Å. The detailed mechanism of ammonia transfer is unknown, but likely involves a conformational rearrangement in the vicinity of a putative channel that connects the two domains (Endrizzi et al., 2004; Goto et al., 2004). One prediction is that a conformational change, induced by UTP and ATP binding, rotates the GATase domain toward the ALase domain to create a shorter channel between the active sites (Goto et al., 2004). Such a large-scale rotation would be unattainable in the steric environment of the filament, as it would lead to clashing of the moving GATase domain with an adjacent CtpS tetramer (Figure 5B,C). Regardless of the specific changes involved, quaternary constraints imposed by the filament structure likely provide the mechanism for inhibition of the synthesis reaction.10.7554/eLife.03638.021Figure 5.Implications of the CtpS filament structure for the mechanism of enzyme inhibition.
Affiliation: Department of Molecular Biology, Princeton University, Princeton, United States.