1T36

Crystal structure of E. coli carbamoyl phosphate synthetase small subunit mutant C248D complexed with uridine 5'-monophosphate

Summary for 1T36

• Entry DOI: 10.2210/pdb1t36/pdb
• Descriptor: Carbamoyl-phosphate synthase large chain, URIDINE-5'-MONOPHOSPHATE, Carbamoyl-phosphate synthase small chain, ... (11 entities in total)
• Functional Keywords: channeling, pyrimidine biosynthesis, arginine biosynthesis, ligase
• Biological source: Escherichia coli; More
• Total number of polymer chains: 8
• Total formula weight: 646617.77

Authors

Thoden, J.B.,Huang, X.,Raushel, F.M.,Holden, H.M. (deposition date: 2004-04-24, release date: 2004-09-21, Last modification date: 2024-10-16)

Primary citation

Thoden, J.B.,Huang, X.,Kim, J.,Raushel, F.M.,Holden, H.M.
Long-range allosteric transitions in carbamoyl phosphate synthetase.
Protein Sci., 13:2398-2405, 2004

PubMed Abstract: Carbamoyl phosphate synthetase plays a key role in both pyrimidine and arginine biosynthesis by catalyzing the production of carbamoyl phosphate from one molecule of bicarbonate, two molecules of MgATP, and one molecule of glutamine. The enzyme from Escherichia coli consists of two polypeptide chains referred to as the small and large subunits, which contain a total of three separate active sites that are connected by an intramolecular tunnel. The small subunit harbors one of these active sites and is responsible for the hydrolysis of glutamine to glutamate and ammonia. The large subunit binds the two required molecules of MgATP and is involved in assembling the final product. Compounds such as L-ornithine, UMP, and IMP allosterically regulate the enzyme. Here, we report the three-dimensional structure of a site-directed mutant protein of carbamoyl phosphate synthetase from E. coli, where Cys 248 in the small subunit was changed to an aspartate. This residue was targeted for a structural investigation because previous studies demonstrated that the partial glutaminase activity of the C248D mutant protein was increased 40-fold relative to the wild-type enzyme, whereas the formation of carbamoyl phosphate using glutamine as a nitrogen source was completely abolished. Remarkably, although Cys 248 in the small subunit is located at approximately 100 A from the allosteric binding pocket in the large subunit, the electron density map clearly revealed the presence of UMP, although this ligand was never included in the purification or crystallization schemes. The manner in which UMP binds to carbamoyl phosphate synthetase is described.

PubMed: 15322282
DOI: 10.1110/ps.04822704

Experimental method

X-RAY DIFFRACTION (2.1 Å)