The conformation of inosine 5'-monophosphate (IMP) bound to IMP dehydrogenase determined by transferred nuclear overhauser effect spectroscopy.
IMP dehydrogenase (IMPDH) catalyzes the NAD-dependent synthesis of xanthosine 5'-monophosphate which is the rate-limiting step in guanine nucleotide biosynthesis. Although IMPDH is the target of numerous chemotherapeutic agents, nothing has been known about the conformation of the enzyme-bound substrates. The conformation of IMP bound to human type II IMP dehydrogenase has been determined by two-dimensional transferred nuclear Overhauser effect NMR spectroscopy at 600 MHz. NOE buildup rates were determined by recording NOESY spectra at numerous mixing times. The cross-relaxation rates determined from the initial NOE build-up rates were used to calculate inter-proton distances of bound IMP. The conformation of the enzyme-bound IMP was obtained by molecular modeling with energy minimization using the experimentally determined inter-proton distance constraints. The glycosidic torsion angle of the bound nucleotide is anti and the sugar is in the C2-endo-conformation. This conformation places H2 of IMP, which is transferred to NAD in the reaction, in a position clear of the rest of the molecule in order to facilitate the reaction.
Xiang
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B
Institute for Cancer Research, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA.
Markham
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GD
D001665:Binding Sites; D006801:Humans; D007168:IMP Dehydrogenase / Q000737:chemistry* / Q000378:metabolism*; D007291:Inosine Monophosphate / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008962:Models, Theoretical; D011485:Protein Binding; D012984:Software; D013816:Thermodynamics
D007291:Inosine Monophosphate; D007168:IMP Dehydrogenase