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-HCI 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 microM for dUMP and N5,10-methylenetetrahydrofolate, respectively) properties determined. 5-Fluoro-dUMP was a competitive, slow-binding inhibitor of the parasite enzyme. N5,10-methylenetetrahydrofolate analogues 10-propargy1-5,8- 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.
Dabrowska
M
M
Nencki Institute of Experimental Biology, Polish Academy of Sciences, Warsaw, Poland.
Zieliñski
Z
Z
Wranicz
M
M
Michalski
R
R
Pawelczak
K
K
Rode
W
W
D000818:Animals; D002846:Chromatography, Affinity; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005468:Fluorodeoxyuridylate / Q000494:pharmacology; D005492:Folic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007700:Kinetics; D007814:Larva; D008099:Liver / Q000201:enzymology; D008115:Liver Regeneration; D009132:Muscles / Q000201:enzymology; D011799:Quinazolines / Q000494:pharmacology; D051381:Rats; D013329:Structure-Activity Relationship; D013876:Thiophenes / Q000494:pharmacology; D013940:Thymidylate Synthase / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D017160:Trichinella spiralis / Q000201:enzymology* / Q000254:growth & development / Q000302:isolation & purification
D004791:Enzyme Inhibitors; D011799:Quinazolines; D013876:Thiophenes; D005468:Fluorodeoxyuridylate; C031662:CB 3717; D005492:Folic Acid; D013940:Thymidylate Synthase; C068874:raltitrexed