The Effects of Removing the GAT Domain from E. coli GMP Synthetase
| Autor: | Gary Lam, Danielle T. Loughlin, Sidney Pollack, Walter A. Patton, Jordan M. Newell, Jessica L. Abbott, Christine M. Lightcap, Melanie A. Weller, Mary E. Olanich |
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| Rok vydání: | 2006 |
| Předmět: |
GMP SYNTHETASE
Protein Conformation Protein domain Bioengineering Molecular cloning Biology medicine.disease_cause Xanthine Biochemistry Analytical Chemistry Domain (software engineering) Adenosine Triphosphate Ammonia Escherichia coli medicine Bioorganic chemistry Carbon-Nitrogen Ligases Cloning Molecular Pyrophosphatases Glutamine amidotransferase Expression vector Escherichia coli Proteins Organic Chemistry Ribonucleotides Recombinant Proteins Protein Structure Tertiary Solutions Kinetics Dimerization |
| Zdroj: | The Protein Journal. 25:483-491 |
| ISSN: | 1875-8355 1572-3887 |
| Popis: | E. coli GMP synthetase (GMPS) catalyzes the conversion of XMP to GMP. Ammonia, generated in the amino-terminal glutamine amidotransferase (GAT) domain, is transferred by an unknown mechanism to the ATP-pyrophosphatase (ATPP) domain, where it attacks a highly reactive adenyl-XMP intermediate, leading to GMP formation. To study the structural requirements for the activity of E. coli GMPS, we used PCR to generate a protein expression construct that contains the ATPP domain as well as the predicted dimerization domain (DD). The ATPP/DD protein is active in solution, utilizing NH (4) (+) as an NH(3) donor. Size-exclusion chromatography demonstrates a dimeric mass for the ATPP/ DD protein, providing the first evidence in solution for the structural organization of the intact GMPS. Kinetic characterization of the ATPP/DD domain protein provides evidence that the presence of the GAT domain can regulate the activity of the ATPP domain. |
| Databáze: | OpenAIRE |
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