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 loss of filamentous mCherry-CtpS localization does not exclude the possibility that the polymerization interface mutants form small filaments that cannot be resolved by light microscopy. Consequently, to determine if the diffuse localization in vivo reflected a polymerization defect, we purified one of the linker region helix mutants, CtpSE277R, and examined its polymerization by light scattering and EM. CtpSE277R did not significantly polymerize in activity buffer, and no filaments could be detected by EM (Figure 7B, Figure 7—figure supplement 1), confirming that CtpSE277R cannot properly polymerize. We attribute the slight linear increase in light scattering with increasing concentration of CtpSE277R to the increase in protein abundance.10.7554/eLife.03638.024Figure 7.Linker helix mutations disrupt polymerization and cause a growth defect.
Affiliation: Department of Molecular Biology, Princeton University, Princeton, United States.