[Comparison of thymidylate synthase properties isolated from the rat tapeworm, hymenolepis diminuta, with properties of enzyme isolated from the host in regenerating rat liver].
Thymidylate synthases (TS) from the tapeworm, Hymenolepis diminuta, and regenerating rat liver have been purified by means of affinity chromatography on immobilized 10-formyl-5,8-dideazafolate and concentrated on immobilized p-aminophenyl-5-fluoro-2'-deoxyuridine monophosphate. Molecular weights of native TS from the tapeworm and regenerating rat liver were 62 kD and 81.5 kD, respectively, and molecular weights of the monomers were 34.4 kD and 34.9 kD, respectively, pointing to dimeric structures of both enzymes. The dependence of TS activity on temperature (Arrhenius plot) was biphasic for the parasite enzyme, with lower activation energy above 32 degrees C, and monophasic for the host enzyme. 2'-deoxyuridine-5'-monophosphate (dUMP) analogues, 5-fluoro-2'-deoxyuridine-5'-monophosphate (5-FdUMP), 2-tio-5-FdUMP,N4-hydroxy-2'-deoxycytidine-5'-monophosphate (N4-hydroxy-dCMP) and N4-hydroxy-5-FdCMP, were competitive with respect to dUMP, slow-binding inhibitors of TS from both sources, with K1 values in 10(-6)-10(-9) M range. 5-FdUMP was distinctly stronger inhibitor of the host than the tapeworm TS, whereas N4-hydroksy-5-FdCMP inhibited stronger the parasite enzyme. Interaction of 5,10-methylenetetrahydrofolate (CH2H4PteGlu) analogue, 10-propargyl-5,8-dideazafolate (pddPteGlu), and its di- and triglutamates with both enzymes were studied. Inhibition of the parasite and host enzymes by pddPteGlu was of mixed-type with respect to CH2H4PteGlu, with K1 values in 10(-8) M range. Introduction of additional glutamate residues changed inhibition type to noncompetitive with respect to Ch2H4PteGlu and lowered K1 values (pddPteGlu3 < pddPteGlu2 < pddPteGlu1).(ABSTRACT TRUNCATED AT 250 WORDS)
Cieśla
J
J
Zakład Biochemii Komórki, Instytut Biologii Doświadczalnej im. M. Nenckiego, Warszawa.
Machnicka
B
B
Pawełczak
K
K
Rzeszotarska
B
B
Rode
I W
IW
D000818:Animals; D006790:Host-Parasite Interactions / Q000502:physiology; D006926:Hymenolepis / Q000502:physiology*; D008099:Liver / Q000201:enzymology*; D008115:Liver Regeneration / Q000502:physiology*; D008970:Molecular Weight; D051381:Rats / Q000469:parasitology*; D013940:Thymidylate Synthase / Q000032:analysis*
D013940:Thymidylate Synthase
Porównanie właściwości syntazy tymidylanowej oczyszczonej z tasiemca szczurzego, Hymenolepis diminuta, z właściwościami enzymu zywiciela wyizolowanego z regenerujacej watroby szczura.