Research Article | Open Access
Jan Massant, Nicolas Glansdorff, "New experimental approaches for investigating interactions between Pyrococcus furiosus carbamate kinase and carbamoyltransferases, enzymes involved in the channeling of thermolabile carbamoyl phosphate", Archaea, vol. 1, Article ID 865962, 9 pages, 2005. https://doi.org/10.1155/2005/865962
New experimental approaches for investigating interactions between Pyrococcus furiosus carbamate kinase and carbamoyltransferases, enzymes involved in the channeling of thermolabile carbamoyl phosphate
A somewhat neglected but essential aspect of the molecular physiology of hyperthermophiles is the protection of thermolabile metabolites and coenzymes. An example is carbamoyl phosphate (CP), a precursor of pyrimidines and arginine, which is an extremely labile and potentially toxic intermediate. The first evidence for a biologically significant interaction between carbamate kinase (CK) and ornithine carbamoyltransferase (OTC) from Pyrococcus furiosus was provided by affinity electrophoresis and co-immunoprecipitation in combination with cross-linking (Massant et al. 2002). Using the yeast two-hybrid system, Hummel-Dreyer chromatography and isothermal titration calorimetry, we obtained additional concrete evidence for an interaction between CK and OTC, the first evidence for an interaction between CK and aspartate carbamoyltransferase (ATC) and an estimate of the binding constant between CK and ATC. The physical interaction between CK and OTC or ATC may prevent thermodenaturation of CP in the aqueous cytoplasmic environment. Here we emphasize the importance of developing experimental approaches to investigate the mechanism of thermal protection of metabolic intermediates by metabolic channeling and the molecular basis of transient protein–protein interactions in the physiology of hyperthermophiles.
- S. Beeckmans, “Chromatographic methods to study protein– protein interactions,” Methods, vol. 19, pp. 278–305, 1999.
- E. Ben-Zeev, A. Berchanski, A. Heifetz, B. Shapira, and M. Eisenstein, “Prediction of the unknown: inspiring experience with the CAPRI experiment,” Proteins, vol. 52, pp. 41–46, 2003.
- C. T. Chien, P. L. Bartel, R. Sternglanz, and S. Fields, “The two-hybrid system: a method to identify and clone genes for proteins that interact with a protein of interest,” Proc. Natl. Acad. Sci. USA, vol. 88, pp. 9578–9582, 1991.
- R. M. Daniel and D. A. Cowan, “Biomolecular stability and life at high temperatures,” Cell. Mol. Life Sci., vol. 57, pp. 250–264, 2000.
- M. L. Doyle, “Characterization of binding interactions by isothermal titration calorimetry,” Curr. Opin. Biotechnol., vol. 8, pp. 31–35, 1997.
- S. Fields and O. Song, “A novel genetic system to detect protein–protein interactions,” Nature, vol. 340, pp. 245–246, 1989.
- S. Fields and R. Sternglanz, “The two-hybrid system: an assay for protein–protein interactions,” Trends Genet., vol. 10, pp. 286–292, 1994.
- J. A. Gegner and F. W. Dahlquist, “Signal transduction in bacteria: CheW forms a reversible complex with the protein kinase CheA,” Proc. Natl. Acad. Sci. USA, vol. 88, pp. 750–754, 1991.
- N. Glansdorff, “Topography of co-transducible arginine mutations in Escherichia coli,” Genetics, vol. 51, pp. 167–179, 1965.
- J. P. Hummel and W. J. Dreyer, “Measurement of protein-binding phenomena by gel filtration,” Biochim. Biophys. Acta, vol. 63, pp. 530–532, 1962.
- P. James, J. Halladay, and E. A. Craig, “Genomic libraries and a host strain designed for highly efficient two-hybrid selection in yeast,” Genetics, vol. 144, pp. 1425–1436, 1996.
- E. Katchalski-Katzir, I. Shariv, M. Eisenstein, A. Friesem, and C. Aflalo, “Molecular surface recognition: determination of geometric fit between proteins and their ligands by correlation techniques,” Proc. Natl. Acad. Sci. USA, vol. 89, pp. 2195–2199, 1992.
- S. Leavitt and E. Freire, “Direct measurement of protein binding energetics by isothermal titration calorimetry,” Curr. Opin. Struct. Biol., vol. 11, pp. 560–566, 2001.
- C. Legrain, M. Demarez, N. Glansdorff, and A. Piérard, “Ammonia-dependent synthesis and metabolic channeling of carbamoyl phosphate in the hyperthermophilic archaeon Pyrococcus furiosus,” Microbiology, vol. 141, pp. 1093–1099, 1995.
- C. Legrain, V. Villeret, and V. Villeret, “Ornithine carbamoyltransferase from Pyrococcus furiosus,” Methods Enzymol., vol. 331, pp. 227–235, 2001.
- B. Li and S. Fields, “Identification of mutations in p53 that affect its binding to SV40 large T antigen by using the yeast two-hybrid system,” FASEB J., vol. 7, pp. 957–963, 1993.
- W. N. Lipscomb, “Aspartate transcarbamylase from Escherichia coli: activity and regulation,” Adv. Enzymol., vol. 68, pp. 67–151, 1994.
- J. Massant, “Molecular physiology of hyperthermophiles: metabolic channeling of carbamoyl phosphate, a thermolabile and potentially toxic intermediate,” Ph.D. Thesis, Vrije Universiteit Brussel, 2004.
- J. Massant and N. Glansdorff, “Metabolic channeling of carbamoyl phosphate in the hyperthermophilic archaeon Pyrococcus furiosus: dynamic enzyme–enzyme interactions involved in the formation of the channeling complex,” Biochem. Soc. Trans., vol. 32, pp. 306–309, 2004.
- J. Massant, P. Verstreken, and P. Verstreken, “Metabolic channeling of carbamoyl phosphate, a thermolabile intermediate: evidence for physical interaction between carbamate kinase-like carbamoyl-phosphate synthetase and ornithine carbamoyltransferase from the hyperthermophile Pyrococcus furiosus,” J. Biol. Chem., vol. 277, pp. 18517–18522, 2002.
- J. Massant, J. Wouters, and N. Glansdorff, “Refined structure of Pyrococcus furiosus ornithine carbamoyltransferase at 1.87Å.,” Acta Crystallogr., vol. D59, pp. 2140–2149, 2003.
- L. McAlister-Henn, N. Gibson, and E. Panisko, “Applications of the yeast two-hybrid system,” Methods, vol. 19, pp. 330–337, 1999.
- E. H. McConkey, “Molecular evolution, intracellular organization, and the quinary structure of proteins,” Proc. Natl. Acad. Sci. USA, vol. 79, pp. 3236–3240, 1982.
- E. M. Phizicky and S. Fields, “Protein–protein interactions: methods for detection and analysis,” Microbiol. Rev., vol. 59, pp. 94–123, 1995.
- M. M. Pierce, C. S. Raman, and B. T. Nall, “Isothermal titration calorimetry of protein–protein interactions,” Methods, vol. 19, pp. 213–221, 1999.
- C. Purcarea, “Aspartate transcarbamoylase from Pyrococcus abyssi,” Methods Enzymol., vol. 331, pp. 248–270, 2001.
- C. Purcarea, D. R. Evans, and G. Hervé, “Channeling of carbamoyl phosphate to the pyrimidine and arginine biosynthetic pathways in the deep sea hyperthermophilic archaeon Pyrococcus abyssi,” J. Biol. Chem., vol. 274, pp. 6122–6129, 1999.
- S. Ramón-Maiques, A. Marina, M. Uriarte, I. Fita, and V. Rubio, “The 1.5 Å resolution crystal structure of the carbamate kinase-like carbamoyl phosphate synthetase from the hyperthermophilic archaeon Pyrococcus furiosus, bound to ADP, confirms that this thermostable enzyme is a carbamate kinase, and provides insight into substrate binding and stability in carbamate kinases.,” J. Mol. Biol., vol. 299, pp. 463–476, 2000.
- M. Uriarte, A. Marina, S. Ramón-Maiques, V. Rubio, V. Durbecq, C. Legrain, and N. Glansdorff, “Carbamoyl phosphate synthesis: carbamate kinase from Pyrococcus furiosus,” Methods Enzymol., vol. 331, pp. 236–247, 2001.
- S. Van Boxstael, R. Cunin, S. Khan, and D. Maes, “Aspartate transcarbamylase from the hyperthermophilic archaeon Pyrococcus abyssi: thermostability and 1.8 Å resolution crystal structure of the catalytic subunit complexed with the bisubstrate analogue N-phosphonacetyl-L-aspartate.,” J. Mol. Biol., vol. 326, pp. 203–216, 2003.
- M. Van de Casteele, C. Legrain, L. Desmarez, P.G. Chen, A. Piérard, and N. Glansdorff, “Molecular physiology of carbamoylation under extreme conditions: what can we learn from extreme thermophilic microorganisms?” Comp. Biochem. Physiol. A, vol. 118, pp. 463–473, 1997.
- V. Villeret, B. Clantin, and B. Clantin, “The crystal structure of Pyrococcus furiosus ornithine carbamoyltransferase reveals a key role for oligomerization in enzyme stability at extremely high temperatures,” Proc. Natl. Acad. Sci. USA, vol. 95, pp. 2801–2806, 1998.
- C. Vélot, M. B. Mixon, M. Teige, and V. A. Srere, “Model of a quinary structure between Krebs TCA cycle enzymes: a model for the metabolon,” Biochemistry, vol. 36, pp. 14271–14276, 1997.
- T. Wiseman, S. Williston, J. F. Brandts, and L.-N. Lin, “Rapid measurement of binding constants and heats of binding using a new titration calorimeter,” Anal. Biochem., vol. 179, pp. 131–137, 1989.
- H. Yong, G. A. Thomas, and W. L. Peticolas, “Metabolite-modulated complex formation between alpha-glycerophosphate dehydrogenase and lactate dehydrogenase,” Biochemistry, vol. 32, pp. 11124–11131, 1993.
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