Characterization of human type I and type II IMP dehydrogenases.
Human IMP dehydrogenase, a target for anticancer and immunosuppressive chemotherapy, exists as two isoforms, types I and II. Nonfusion sequences of each isoform were overexpressed in an IMP dehydrogenase-deficient strain of Escherichia coli and purified to homogeneity. Both recombinant isoforms were tetramers, which was in agreement with the subunit structure of the native mammalian enzyme. The results of initial velocity and product inhibition studies were consistent with an Ordered Bi Bi kinetic mechanism for both isoforms. Substrate affinities were similar for types I and II with Km values of 18 and 9.3 microM, respectively, for IMP, and 46 and 32 microM, respectively, for NAD.kcat values were 1.5 and 1.3 s-1 at 37 degrees C for types I and II, respectively. Xanthosine 5'-monophosphate and NADH inhibited the two isoforms with identical inhibition patterns and inhibition constants. Mycophenolic acid, however, inhibited the type II enzyme with a 4.8-fold lower K than the type I. Selective inhibitors of the inducible type II isoform may mitigate toxicity caused by inhibition of the constitutively expressed type I isoform.
Carr
S F
SF
Institute of Biochemistry and Cell Biology, Syntex Discovery Research, Palo Alto, California 94303.
Papp
E
E
Wu
J C
JC
Natsumeda
Y
Y
D000595:Amino Acid Sequence; D002384:Catalysis; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D006801:Humans; D007168:IMP Dehydrogenase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity
D007527:Isoenzymes; D011994:Recombinant Proteins; D007168:IMP Dehydrogenase