Orotate phosphoribosyltransferase and orotidine 5'-monophosphate decarboxylase exist as multienzyme complex in human malaria parasite Plasmodium falciparum.
Plasmodium falciparum, the causative agent of the most lethal form of human malaria, totally depends on de novo pyrimidine biosynthetic pathway. Orotate phosphoribosyltransferase (OPRT) and orotidine 5'-monophosphate decarboxylase (OMPDC), the fifth and sixth enzymes in the pathway catalyzing formation of uridine 5'-monophosphate (UMP), remain largely uncharacterized in the protozoan parasite. In this study, we achieved purification of OPRT and OMPDC to near homogeneity from P. falciparum cultivated in vitro. The OPRT and OMPDC activities were co-eluted in all chromatographic columns during purification, suggesting the purified proteins exist as a multienzyme complex with a molecular mass of 140+/-8 kDa and contain two subunits each of OPRT and OMPDC. Monomeric forms of OPRT and OMPDC had molecular masses of 32+/-3 and 38+/-3 kDa, respectively, in agreement with those of proteins predicted from P. falciparum genome database. Interestingly, kinetic parameters and inhibitory constants of both OPRT and OMPDC activities were found to be different to those of the bifunctional human red cell UMP synthase. Our evidence provides the first example of OPRT and OMPDC existing as a multienzyme complex.
Krungkrai
Sudaratana R
SR
Department of Biochemistry, Faculty of Medicine, Chulalongkorn University, Rama 4 Road, Bangkok 10330, Thailand.
Prapunwattana
Phisit
P
Horii
Toshihiro
T
Krungkrai
Jerapan
J
D000595:Amino Acid Sequence; D000818:Animals; D002850:Chromatography, Gel; D002853:Chromatography, Liquid / Q000379:methods; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009097:Multienzyme Complexes / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D009962:Orotate Phosphoribosyltransferase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D009964:Orotidine-5'-Phosphate Decarboxylase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D010962:Plasmodium berghei / Q000201:enzymology; D010963:Plasmodium falciparum / Q000201:enzymology*; D021122:Protein Subunits; D014542:Uridine Monophosphate / Q000031:analogs & derivatives / Q000378:metabolism
D009097:Multienzyme Complexes; D021122:Protein Subunits; D014542:Uridine Monophosphate; D009962:Orotate Phosphoribosyltransferase; D009964:Orotidine-5'-Phosphate Decarboxylase