Large-scale filament formation inhibits the activity of CTP synthetase
| Autor: | Jesse M Hansen, Emeric J. Charles, Enoch P. Baldwin, Hsin Jung Li, Alexander Lorestani, Anne-Florence Bitbol, Chris H. Habrian, Rachael M. Barry, Zemer Gitai, Justin M. Kollman, Ned S. Wingreen |
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| Jazyk: | angličtina |
| Rok vydání: | 2014 |
| Předmět: |
Models
Molecular Conformational change Mutant Gene Expression Biochemistry chemistry.chemical_compound Models Site-Directed Carbon-Nitrogen Ligases CTP synthetase Biology (General) chemistry.chemical_classification biology Fluid Membranes General Neuroscience Escherichia coli Proteins General Medicine Cytoophidium Medicine enzyme regulation nucelotide metabolism Research Article Cytidine triphosphate QH301-705.5 Science Cytidine Triphosphate Recombinant Fusion Proteins General Biochemistry Genetics and Molecular Biology Escherichia coli General Immunology and Microbiology Prevention Mutagenesis E. coli Molecular Cell Biology pyrimidine metabolism Kinetics Enzyme chemistry Polymerization biology.protein Biophysics Mutagenesis Site-Directed Biochemistry and Cell Biology Protein Multimerization |
| Zdroj: | Barry, RM; Bitbol, AF; Lorestani, A; Charles, EJ; Habrian, CH; Hansen, JM; et al.(2014). Large-scale filament formation inhibits the activity of CTP synthetase. eLife, 3(July2014), 1-19. doi: 10.7554/eLife.03638. UC Davis: Retrieved from: http://www.escholarship.org/uc/item/1tc7h0fp eLife eLife, vol 3, iss July2014 eLife, Vol 3 (2014) |
| Popis: | 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. DOI: http://dx.doi.org/10.7554/eLife.03638.001 eLife digest Enzymes are proteins that perform reactions that can convert one or more chemicals (the substrates) into others (the products). The rate at which an enzyme produces its product is often carefully regulated. Some molecules slow or stop an enzyme by binding to and blocking the site where its substrates normally bind: its ‘active site’. Other molecules can also bind to sites other than the active site, which can cause the enzyme to become either more or less active. Almost all living things have an enzyme called CTP synthetase that makes one of the building blocks that is used to build DNA and a similar molecule called RNA. This enzyme converts a molecule called uridine triphosphate (or UTP) into another called cytidine triphosphate (CTP): a reaction that is powered by breaking down molecules of adenosine triphosphate (ATP). The amount of CTP synthetase made by a cell is carefully controlled. The enzyme's activity is also regulated by the levels of UTP and CTP, and by another molecule (called GTP) that binds to a site outside of its active site. Four copies of the CTP synthetase protein must work together before this enzyme can turn UTP into CTP. The enzyme also forms much larger aggregates, or polymers; however, it is not clear what causes these polymers to form, or what they do in a cell. Barry et al. have now discovered that CTP synthetase is almost completely inactivated when these polymers are formed. Furthermore, CTP encourages the polymers to form, whilst UTP and ATP cause them to disassemble. Therefore, this enzyme is least active when there is excess product in the cell, and most active when its substrates are plentiful. By determining the three-dimensional structure of a CTP synthetase polymer, Barry et al. reveal that although CTP is bound to the enzymes, their active sites are still freely accessible. However, the enzymes in the polymer appear to be locked into a shape that makes them unable to carry out their function. When Barry et al. then mutated the enzyme so that it was unable to form polymers it was also no longer inactivated in the same way by CTP. Bacterial cells with only these mutant versions of CTP synthetase are unable to properly control their levels of CTP. This suggests that polymer formation is important for regulating this enzyme in response to a build up of its product. Further work is needed to see whether the regulation of CTP synthetase activity by forming polymers is specific to this enzyme or a widespread mechanism that is used to control other enzymes too. DOI: http://dx.doi.org/10.7554/eLife.03638.002 |
| Databáze: | OpenAIRE |
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