Structure and inhibition of orotidine 5'-monophosphate decarboxylase from Plasmodium falciparum.
Orotidine 5'-monophosphate (OMP) decarboxylase from Plasmodium falciparum (PfODCase, EC 4.1.1.23) has been overexpressed, purified, subjected to kinetic and biochemical analysis, and crystallized. The native enzyme is a homodimer with a subunit molecular mass of 38 kDa. The saturation curve for OMP as a substrate conformed to Michaelis-Menten kinetics with K m = 350 +/- 60 nM and V max = 2.70 +/- 0.10 micromol/min/mg protein. Inhibition patterns for nucleoside 5'-monophosphate analogues were linear competitive with respect to OMP with a decreasing potency of inhibition of PfODCase in the order: pyrazofurin 5'-monophosphate ( K i = 3.6 +/- 0.7 nM) > xanthosine 5'-monophosphate (XMP, K i = 4.4 +/- 0.7 nM) > 6-azauridine 5'-monophosphate (AzaUMP, K i = 12 +/- 3 nM) > allopurinol-3-riboside 5'-monophosphate ( K i = 240 +/- 20 nM). XMP is an approximately 150-fold more potent inhibitor of PfODCase compared with the human enzyme. The structure of PfODCase was solved in the absence of ligand and displays a classic TIM-barrel fold characteristic of the enzyme. Both the phosphate-binding loop and the betaalpha5-loop have conformational flexibility, which may be associated with substrate capture and product release along the reaction pathway.
Langley
David B
DB
School of Molecular and Microbial Biosciences, University of Sydney, Sydney, New South Wales 2006, Australia.
Shojaei
Maryam
M
Chan
Camilla
C
Lok
Hiu Chuen
HC
Mackay
Joel P
JP
Traut
Thomas W
TW
Guss
J Mitchell
JM
Christopherson
Richard I
RI
D000818:Animals; D001665:Binding Sites; D003460:Crystallization; D018360:Crystallography, X-Ray; D019281:Dimerization; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D008970:Molecular Weight; D009964:Orotidine-5'-Phosphate Decarboxylase / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D010963:Plasmodium falciparum / Q000201:enzymology; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry; D012265:Ribonucleotides / Q000494:pharmacology; D013045:Species Specificity; D014542:Uridine Monophosphate / Q000031:analogs & derivatives / Q000494:pharmacology
D011994:Recombinant Proteins; D012265:Ribonucleotides; C041404:allopurinol riboside 5'-monophosphate; C028372:1-(5'-phospho-beta-D-ribofuranosyl)barbituric acid; D014542:Uridine Monophosphate; D009964:Orotidine-5'-Phosphate Decarboxylase