Conformational Changes and Stabilization of Inosine 5′-Monophosphate Dehydrogenase Associated with Ligand Binding and Inhibition by Mycophenolic Acid

Nimmesgern, Elmar; Fox, Ted; Fleming, Mark; Thomson, John A.

doi:10.1074/jbc.271.32.19421

Cited by 47 publications

(37 citation statements)

References 46 publications

(54 reference statements)

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“…Further, the presence of the SAD dinucleotide analogue may partially order the end of the active site flap, suggesting that the flap may stabilize both substrate and cofactor binding in the normal ternary complex. This is consistent with the observation that substrateinduced protection of the flap against proteolysis is enhanced by cofactor binding (66).…”

Section: Summary and Discussionsupporting

confidence: 80%

Crystal structure of human type II inosine monophosphate dehydrogenase: Implications for ligand binding and drug design

Colby

Vanderveen

Strickler

et al. 1999

Proc. Natl. Acad. Sci. U.S.A.

138

127

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Inosine monophosphate dehydrogenase (IM-PDH) controls a key metabolic step in the regulation of cell growth and differentiation. This step is the NAD-dependent oxidation of inosine 5 monophosphate (IMP) to xanthosine 5 monophosphate, the rate-limiting step in the synthesis of the guanine nucleotides. Two isoforms of IMPDH have been identified, one of which (type II) is significantly up-regulated in neoplastic and differentiating cells. As such, it has been identified as a major target in antitumor and immunosuppressive drug design. We present here the 2.9-Å structure of a ternary complex of the human type II isoform of IMPDH. The complex contains the substrate analogue 6-chloropurine riboside 5-monophosphate (6-Cl-IMP) and the NAD analogue selenazole-4-carboxamide adenine dinucleotide, the selenium derivative of the active metabolite of the antitumor drug tiazofurin. The enzyme forms a homotetramer, with the dinucleotide binding at the monomer-monomer interface. The 6 chloro-substituted purine base is dehalogenated, forming a covalent adduct at C6 with Cys-331. The dinucleotide selenazole base is stacked against the 6-Cl-IMP purine ring in an orientation consistent with the B-side stereochemistry of hydride transfer seen with NAD. The adenosine end of the ligand interacts with residues not conserved between the type I and type II isoforms, suggesting strategies for the design of isoform-specific agents.

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Section: Summary and Discussionsupporting

confidence: 80%

Crystal structure of human type II inosine monophosphate dehydrogenase: Implications for ligand binding and drug design

Colby

Vanderveen

Strickler

et al. 1999

Proc. Natl. Acad. Sci. U.S.A.

138

127

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“…Treatment with MPA results in the formation of large aggregates with a tendency to annular formation, although without the same level of definition found in intact cells. These data demonstrate that MPA causes self aggregation of IMPDH in the absence of other proteins or cellular elements, most likely through conformational change (20).…”

Section: Resultsmentioning

confidence: 88%

Regulation of the Interaction of Inosine Monophosphate Dehydrogenase with Mycophenolic Acid by GTP

Makhov

et al. 2006

Journal of Biological Chemistry

110

127

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Inosine monophosphate dehydrogenase (IMPDH), a rate-limiting enzyme in the de novo synthesis of guanine nucleotides, is a major therapeutic target. A prototypic uncompetitive inhibitor of IMPDH, mycophenolic acid (MPA), is the active form of mycophenolate mofeteil (CellCept), a widely used immunosuppressive drug. We have found that MPA interacts with intracellular IMPDH in vivo to alter its mobility on SDS-polyacrylamide gels. MPA also induces a striking conformational change in IMPDH protein in intact cells, resulting in the formation of annular aggregates of protein with concomitant inhibition of IMPDH activity. These aggregates are not associated with any known intracellular organelles and are reversible by incubating cells with guanosine, which repletes intracellular GTP, or with GTP␥S. GTP also restores IMPDH activity. Treatment of highly purified IMPDH with MPA also results in the formation of large aggregates of protein, a process that is both prevented and reversed by the addition of GTP. Finally, GTP binds to IMPDH at physiologic concentrations, induces the formation of linear arrays of tetrameric protein, and prevents the aggregation of protein induced by MPA. We conclude that intracellular GTP acts as an antagonist to MPA by directly binding to IMPDH and reversing the conformational changes in the protein. Inosine monophosphate dehydrogenase (IMPDH)3 catalyzes the first step in the de novo synthetic pathway for the formation of guanine nucleotides by converting IMP to xanthosine 5Ј-monophosphate (XMP) with the concomitant reduction of NAD ϩ . As the rate-limiting step in this pathway, the enzyme has been identified as an important regulator of cell proliferation. Inhibitors of IMPDH are in clinical use as immunosuppressive agents and have potential utility in the treatment of neoplastic and viral diseases (1-5). Two isoforms of IMPDH have been identified, the type I form that is expressed at lower levels in all cell types and the type II form that is more highly expressed in proliferating and transformed cell types (6, 7). These enzymes have 84% amino acid identity, and both are catalytically active as tetramers of 55-kDa subunits with similar, although not identical, kinetic profiles. Studies on the regulation of IMPDH activity have demonstrated strong transcriptional up-regulation of the type II mRNA in response to mitogenic stimuli in peripheral blood lymphocytes and in response to enzyme inhibition and guanine nucleotide depletion in a number of cell types (8, 9). Until recently, very little attention has been paid to the role of protein structure in regulating its activity. The finding that mutations within the IMPDH type I coding region are associated with a familial form of retinitis pigmentosa (10) has initiated renewed interest in the structure of IMPDH, first crystallized in 1996 (11). These mutations occur within a region of the protein termed the cystathione ␤-synthase domain that has recently been shown to bind oligonucleotides up to 100-base pair in length (12). In addition, this domain ...

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“…26 Later studies showed that binding of both IMP and NAD ϩ were required to induce a conformational rearrangement to produce a closed form of the enzyme. 27 After IMP and NAD ϩ are bound, IMP is oxidized to XMP*, a covalently bound intermediate. Once NADH leaves, MPA binds to form an inhibited complex with nanomolar affinity.…”

Section: Ligand Solution Structures By Transferred Noe Methodsmentioning

confidence: 99%

NMR techniques for characterization of ligand binding: Utility for lead generation and optimization in drug discovery

Moore

1999

Biopolymers

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Over the last ten years, nmr spectroscopy has evolved into an important discipline in drug discovery. Initially, nmr was most useful as a technique to provide structural information regarding protein drug targets and target–ligand interactions. More recently, it has been shown that nmr may be used as an alternative method for identification of small molecule ligands that bind to protein drug targets. High throughput implementation of these experiments to screen small molecule libraries may lead to identification of potent and novel lead compounds. In this review, we will use examples from our own research to illustrate how nmr experiments to characterize ligand binding may be used to both screen for novel compounds during the process of lead generation, as well as provide structural information useful for lead optimization during the latter stages of a discovery program. © 1999 John Wiley & Sons, Inc. Biopoly 51: 221–243, 1999

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Conformational Changes and Stabilization of Inosine 5′-Monophosphate Dehydrogenase Associated with Ligand Binding and Inhibition by Mycophenolic Acid

Cited by 47 publications

References 46 publications

Crystal structure of human type II inosine monophosphate dehydrogenase: Implications for ligand binding and drug design

Crystal structure of human type II inosine monophosphate dehydrogenase: Implications for ligand binding and drug design

Regulation of the Interaction of Inosine Monophosphate Dehydrogenase with Mycophenolic Acid by GTP

NMR techniques for characterization of ligand binding: Utility for lead generation and optimization in drug discovery

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