Large-scale Filament Formation Inhibits the Activity of CTP Synthetase

Elife. 2014 Jul 16;3:e03638. doi: 10.7554/eLife.03638.

Abstract

CTP Synthetase (CtpS) is a universally conserved and essential metabolic enzyme. While many enzymes form small oligomers, CtpS forms large-scale filamentous structures of unknown function in prokaryotes and eukaryotes. By simultaneously monitoring CtpS polymerization and enzymatic activity, we show that polymerization inhibits activity, and CtpS's product, CTP, induces assembly. To understand how assembly inhibits activity, we used electron microscopy to define the structure of CtpS polymers. This structure suggests that polymerization sterically hinders a conformational change necessary for CtpS activity. 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.

Keywords: CTP synthetase; enzyme regulation; nucelotide metabolism; pyrimidine metabolism.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Carbon-Nitrogen Ligases / chemistry
  • Carbon-Nitrogen Ligases / genetics
  • Carbon-Nitrogen Ligases / metabolism*
  • Cytidine Triphosphate / biosynthesis*
  • Escherichia coli / chemistry
  • Escherichia coli / enzymology*
  • Escherichia coli / genetics
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / genetics
  • Escherichia coli Proteins / metabolism*
  • Gene Expression
  • Kinetics
  • Models, Molecular
  • Mutagenesis, Site-Directed
  • Protein Multimerization
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism*

Substances

  • Escherichia coli Proteins
  • Recombinant Fusion Proteins
  • Cytidine Triphosphate
  • Carbon-Nitrogen Ligases
  • CTP synthetase