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) |
|---|---|
| Pages (from-to) | 483-491 |
| Number of pages | 9 |
| Journal | Protein Journal |
| Volume | 25 |
| Issue number | 7-8 |
| DOIs | |
| State | Published - Dec 1 2006 |
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All Science Journal Classification (ASJC) codes
- Analytical Chemistry
- Bioengineering
- Biochemistry
- Organic Chemistry
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The effects of removing the GAT domain from E. coli GMP synthetase. / Abbott, Jessica L.; Newell, Jordan; Lightcap, Christine M.; Olanich, Mary E.; Loughlin, Danielle T.; Weller, Melanie A.; Lam, Gary; Pollack, Sidney; Patton, Walter A.
In: Protein Journal, Vol. 25, No. 7-8, 01.12.2006, p. 483-491.Research output: Contribution to journal › Article
TY - JOUR
T1 - The effects of removing the GAT domain from E. coli GMP synthetase
AU - Abbott, Jessica L.
AU - Newell, Jordan
AU - Lightcap, Christine M.
AU - Olanich, Mary E.
AU - Loughlin, Danielle T.
AU - Weller, Melanie A.
AU - Lam, Gary
AU - Pollack, Sidney
AU - Patton, Walter A.
PY - 2006/12/1
Y1 - 2006/12/1
N2 - 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.
AB - 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.
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UR - http://www.scopus.com/inward/citedby.url?scp=33845577927&partnerID=8YFLogxK
U2 - 10.1007/s10930-006-9032-5
DO - 10.1007/s10930-006-9032-5
M3 - Article
VL - 25
SP - 483
EP - 491
JO - Protein Journal
JF - Protein Journal
SN - 1572-3887
IS - 7-8
ER -