Ornithine decarboxylase from Leishmania mexicana promastigotes: interaction with pyridoxal 5'-phosphate and alpha-difluoromethylornithine.
The catalytic properties of ornithine decarboxylase (ODC) from Leishmania mexicana as well as the interaction with its cofactor pyridoxal 5'-phosphate (PLP) and the irreversible inhibitor alpha-difluoromethylornithine (DFMO) have been studied using partially purified preparations of the enzyme obtained from parasite promastigotes. Leishmania extracts prepared in the presence of saturating concentrations of PLP yielded an enzyme considerably more resistant to heat inactivation and with a three-fold higher activity than the ODC obtained without the addition of cofactor. The complete removal of PLP by treatment with hydroxylamine yielded the apoenzyme which shows an absolute requirement for PLP to recover its enzymatic activity. The Km values for L-ornithine and PLP were 0.7 mM and 25 microM, respectively, while Ki for DFMO was 0.2 mM. The restoration of ODC activity from apoenzyme and cofactor seems to involve time and temperature-dependent activation processes. L. mexicana ODC has an apparent molecular mass of 240 +/- 20 kDa.
Sánchez
C P
CP
Instituto de Investigaciones Bioquímicas "Fundación Campomar", University of Buenos Aires, Argentina.
Sidrauski
C
C
Freire
S M
SM
González
N S
NS
Algranati
I D
ID
D000818:Animals; D001051:Apoenzymes / Q000378:metabolism; D002384:Catalysis; D002850:Chromatography, Gel; D000518:Eflornithine / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D004793:Enzyme Reactivators / Q000494:pharmacology; D006358:Hot Temperature; D019811:Hydroxylamine; D006898:Hydroxylamines / Q000494:pharmacology; D007894:Leishmania mexicana / Q000201:enzymology*; D008970:Molecular Weight; D009955:Ornithine Decarboxylase / Q000737:chemistry / Q000378:metabolism*; D065108:Ornithine Decarboxylase Inhibitors; D011732:Pyridoxal Phosphate / Q000494:pharmacology*
D001051:Apoenzymes; D004793:Enzyme Reactivators; D006898:Hydroxylamines; D065108:Ornithine Decarboxylase Inhibitors; D019811:Hydroxylamine; D011732:Pyridoxal Phosphate; D009955:Ornithine Decarboxylase; D000518:Eflornithine