PMID:17103135
| Citation |
Abbott, JL, Newell, JM, Lightcap, CM, Olanich, ME, Loughlin, DT, Weller, MA, Lam, G, Pollack, S and Patton, WA (2006) The effects of removing the GAT domain from E. coli GMP synthetase. Protein J. 25:483-91 |
|---|---|
| Abstract |
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. |
| Links |
PubMed Online version:10.1007/s10930-006-9032-5 |
| Keywords |
Adenosine Triphosphate/chemistry; Ammonia/chemistry; Carbon-Nitrogen Ligases/chemistry; Carbon-Nitrogen Ligases/genetics; Carbon-Nitrogen Ligases/isolation & purification; Cloning, Molecular; Dimerization; Escherichia coli/enzymology; Escherichia coli Proteins/chemistry; Kinetics; Protein Conformation; Protein Structure, Tertiary; Pyrophosphatases/chemistry; Pyrophosphatases/genetics; Pyrophosphatases/isolation & purification; Recombinant Proteins/chemistry; Recombinant Proteins/isolation & purification; Ribonucleotides/chemistry; Solutions |
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