F. Hidalgo-zarco et al., Kinetic properties and inhibition of the dimeric dUTPase-dUDPase from Leishmania major, PROTEIN SCI, 10(7), 2001, pp. 1426-1433
Kinetic properties of the dimeric enzyme dUTPase from Leishmania major were
studied using a continuous spectrophotometric method. dUTP was the natural
substrate and dUMP and PPi the products of the hydrolysis. The trypanosoma
tid enzyme exhibited a low K-m value for dUTP (2.11 muM), a k(cat) of 49 s(
-1), strict Michaelis-Menten kinetics and is a potent catalyst of dUDP hydr
olysis, whereas in other dUTPases described, this compound acts as a compet
itive inhibitor. Discrimination is achieved for the base and sugar moiety s
howing specificity constants for different dNTPs similar to those of bacter
ial, viral, and human enzymes. In the alkaline range, the K-m for dUTP incr
eases with the dissociation of ionizable groups showing pK(a) values of 8.8
, identified as the uracil moiety of dUTP and 10, whereas in the acidic ran
ge, K-m is regulated by an enzyme residue exhibiting a pK(a) of 7.1. Activi
ty is strongly inhibited by the nucleoside triphosphate analog alpha-beta -
imido-dUTP, indicating that the enzyme can bind triphosphate analogs. The e
xistence of specific inhibition and the apparent structural and kinetic dif
ferences (reflected in different binding strength of dNTPs) with other euka
ryotic dUTPases suggest that the present enzyme might be exploited as a tar
get for new drugs against leishmaniasis.