Identification of a folate binder in hog kidney.
A macromolecular binder of folic acid (pteroylglutamic acid) and folic acid derivatives has been identified in extracts of hog kidney. With partially purified preparations, binding of [3H]pteroylglutamate was competed for by unlabeled pteroylglutamate, 5-methyltetrahydrofolic acid and its triglutamate derivative, by tetra- and dihydrofolic acid, and by N-10-formyltetrahydrofolic acid. The partially purified extract did not bine [3H]methotrexate nor could methotrexate or 5-formyltetrahydrofolic acid compete for [3H]folic acid-binding sites. The rate of binding of pterolyglutamate at 37 degrees was approximately 3%/s, was independent of pteroylglutamate concentration, and was essentially irreversible between pH 6.0 and 9.0. Below pH 6.0 binding was reversible, and at pH 3.5 the folic acid-binder complex completely disassociated. Based upon Sephadex gel filtration, the molecular weight of the folate-binder complex is 35,000 to 40,000. Binding activity was unaffected by pretreatment with ribonuclease or deoxyribonuclease but was completely destroyed by trypsin. The initial, unfractionated extract showed gamma-glutamyl carboxypeptidase (conjugase) activity which was lost in subsequent steps of purification of the folate binder.
D000818:Animals; D001665:Binding Sites; D002268:Carboxypeptidases; D005492:Folic Acid / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000378:metabolism*; D008727:Methotrexate / Q000378:metabolism; D011485:Protein Binding; D013552:Swine; D013763:Tetrahydrofolates / Q000378:metabolism; D014357:Trypsin
D013763:Tetrahydrofolates; D005492:Folic Acid; D002268:Carboxypeptidases; D014357:Trypsin; D008727:Methotrexate