Ins. Subbayya et H. Balaram, Evidence for multiple active states of Plasmodium falciparum hypoxanthine-guanine-xanthine phosphoribosyltransferase, BIOC BIOP R, 279(2), 2000, pp. 433-437
Citations number
18
Categorie Soggetti
Biochemistry & Biophysics
Journal title
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
The lack of de novo purine biosynthesis in the malaria parasite Plasmodium
falciparum makes the purine salvage enzyme hypoxanthine-guanine-xanthine ph
osphoribosyltransferase (HGXPRT) a drug target. However, high activities fo
r the purified recombinant enzyme have not been achieved, indicating that t
he P. falciparum enzyme requires very precise conditions for its maximal ac
tivity. In this report we have standardized the activation conditions neces
sary for high levels of activity, which is critically dependent on the rati
os of enzyme, phosphoribosylpyrophosphate (PRPP), hypoxanthine, and buffer
conditions. We demonstrate that excess substrates will! push the enzyme to
a less active state. We also present evidence for the existence of differen
t kinetic states of the enzyme during activation and storage. Our kinetic d
ata show that hypoxanthine is the substrate with highest affinity for the e
nzyme with a K-m well below 1 muM. The activated enzyme has a maximum activ
ity of 8.370 mu mol/min/mg for hypoxanthine which is 10.8 times more than t
he previous reports. We discuss the biological relevance and implications o
f these results on drug design efforts. (C) 2000 Academic Press.