title: Dissociation of the octameric bifunctional enzyme formiminotransferase-cyclodeaminase in urea. Isolation of two monofunctional dimers.
abstract: Partial denaturation of the circular octameric bifunctional enzyme formiminotransferase-cyclodeaminase in increasing urea concentrations leads to sequential dissociation via dimers to inactive monomers. In potassium phosphate buffer, dissociation to dimers in 3 M urea coincides with loss of both activities and a major decrease in intensity of intrinsic tryptophan fluorescence. In the presence of folic acid, these dimers retain the deaminase activity, but with folylpolyglutamates, both activities are protected and the native octameric structure is retained. The protection profiles with polyglutamates are cooperative with a Hill coefficient greater than 2, suggesting that binding of more than one folylpolyglutamate per octamer is required to stabilize the native structure. In triethanolamine hydrochloride buffer, transferase-active dimers that retain the intrinsic tryptophan fluorescence can be obtained in 1 M urea and stabilized at higher urea concentration by the addition of glutamate. Deaminase-active dimers are obtained by the protection of folate in 3 M urea. Proteolysis of the two kinds of dimers by chymotrypsin leads to very different fragmentation patterns, indicating that they are structurally different. We propose that the two dimers retain different subunit-subunit interfaces, one of which is required for each activity. This suggests that the native octameric structure is required for expression of both activities and therefore for "channeling" of intermediates.
mesh_terms: D000642:Ammonia-Lyases / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D011489:Protein Denaturation; D011624:Pteroylpolyglutamic Acids / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D014508:Urea / Q000494:pharmacology*
chemical_list: D046911:Macromolecular Substances; D011624:Pteroylpolyglutamic Acids; D014508:Urea; D000642:Ammonia-Lyases; C020148:formiminotetrahydrofolate cyclodeaminase