Allosteric control of the oligomerization of carbamoyl phosphate synthetase from Escherichia coli

J. Kim, F. M. Raushel

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

Carbamoyl phosphate synthetase (CPS) from Escherichia coli is allosterically regulated by the metabolites ornithine, IMP, and UMP. Ornithine and IMP function as activators, whereas UMP is an inhibitor. CPS undergoes changes in the state of oligomerization that are dependent on the protein concentration and the binding of allosteric effectors. Ornithine and IMP promote the formation of an (αβ)4 tetramer while UMP favors the formation of an (αβ)2 dimer. The three-dimensional structure of the (αβ)4 tetramer has unveiled two regions of molecular contact between symmetry-related monomeric units. Identical residues within two pairs of allosteric domains interact with one another as do twin pairs of oligomerization domains. There are thus two possible structures for an (αβ)2 dimer: an elongated dimer formed at the interface of two allosteric domains and a more compact dimer formed at the interface between two oligomerization domains. Mutations at the two interfacial sites of oligomerization were constructed in an attempt to elucidate the mechanism for assembly of the (αβ)4 tetramer through disruption of the molecular binding interactions between monomeric units. When Leu-421 (located in the oligomerization domain) was mutated to a glutamate residue, CPS formed an (αβ)2 dimer in the presence of ornithine, UMP, or IMP. In contrast, when Asn-987 (located in the allosteric binding domain) was mutated to an aspartate, an (αβ) monomer was formed regardless of the presence of any allosteric effectors. These results are consistent with a model for the structure of the (αβ)2 dimer that is formed through molecular contact between two pairs of allosteric domains. Apparently, the second interaction, between pairs of oligomerization domains, does not form until after the interaction between pairs of allosteric domains is formed. The binding of UMP to the allosteric domain inhibits the dimerization of the (αβ)2 dimer, whereas the binding of either IMP or ornithine to this same domain promotes the dimerization of the (αβ)2 dimer. In the oligomerization process, ornithine and IMP must exert a conformational alteration on the oligomerization domain, which is ∼45 Å away from their site of binding within the allosteric domain. No significant dependence of the specific catalytic activity on the protein concentration could be detected, and thus the effects induced by the allosteric ligands on the catalytic activity and the state of oligomerization are unlinked from one another.

Original languageEnglish (US)
Pages (from-to)11030-11036
Number of pages7
JournalBiochemistry
Volume40
Issue number37
DOIs
StatePublished - Sep 18 2001
Externally publishedYes

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Carbamyl Phosphate
Inosine Monophosphate
Oligomerization
Ornithine
Uridine Monophosphate
Ligases
Escherichia coli
Dimers
Dimerization
Activator Appliances
Catalyst activity
Protein Binding
Aspartic Acid
Glutamic Acid
Binding Sites
Metabolites
Ligands
Mutation
Proteins
Monomers

ASJC Scopus subject areas

  • Biochemistry

Cite this

Allosteric control of the oligomerization of carbamoyl phosphate synthetase from Escherichia coli. / Kim, J.; Raushel, F. M.

In: Biochemistry, Vol. 40, No. 37, 18.09.2001, p. 11030-11036.

Research output: Contribution to journalArticle

Kim, J. ; Raushel, F. M. / Allosteric control of the oligomerization of carbamoyl phosphate synthetase from Escherichia coli. In: Biochemistry. 2001 ; Vol. 40, No. 37. pp. 11030-11036.
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