The interaction of pteroylpolyglutamates with calf thymus thymidylate synthase

Calf thymus thymidylate synthase was purified to homogeneity using a pteroyltetraglutamyllysine Sepharose affinity column. Inhibition of the purified enzyme by folate or methotrexate was enhanced by addition of gamma-Glu residues. Tetrahydrofolate, tetrahydropteroyltriglutamate and tetrahydropteroylheptaglutamate, all having the natural configuration at C-6, all showed similar Km values near 15 microM. Thus, although calf thymus thymidylate synthase showed a higher affinity for pteroylpolyglutamates than pteroylmonoglutamate, this was not reflected in lowered Km values with the corresponding tetrahydropteroylpolyglutamate substrates.