Trichinella spiralis thymidylate synthase: Developmental pattern, isolation, molecular properties, and inhibition by substrate and cofactor analogues

Thymidylate synthase specific activity was found to remain at a constant level in crude extracts from muscle larvae, isolated (1-15 months after infection) by pepsin-HCl digestion, as well as from adult worms of Trichinella spiralis. The enzyme was purified and its molecular (monomer mol. wt 35 kD) and kinetic (sequential mechanism with the K-m values 3.1 and 19 mu M for dUMP and N-5,N-10-methylenetetrahydrofolate, respectively) properties determined. 5-Fluoro-dUMP was a competitive, slow-binding inhibitor of the parasite enzyme. N-5,N-10-methylenetetrahydrofolate analogues 10-propargyl-5,8-dideazafolate (CB3717), ZD1694, BW1843U89, and AG337 were weaker inhibitors of the parasite than regenerating rat liver enzyme. Inhibition by 10-propargyl-5,8-dideazafolate was strengthened by an increasing number of glutamate residues. Thymidine kinase activity could not be detected in the muscle larvae crude extracts. (C) 1996 Academic Press, Inc.