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 NH3 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.
Original language | English (US) |
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Pages (from-to) | 483-491 |
Number of pages | 9 |
Journal | Protein Journal |
Volume | 25 |
Issue number | 7-8 |
DOIs | |
State | Published - Dec 2006 |
All Science Journal Classification (ASJC) codes
- Analytical Chemistry
- Bioengineering
- Biochemistry
- Organic Chemistry