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Nucleotide binding to human UMP-CMP kinase using fluorescent derivatives - a screening based on affinity for the UMP-CMP binding site.

Abstract : Methylanthraniloyl derivatives of ATP and CDP were used in vitro as fluorescent probes for the donor-binding and acceptor-binding sites of human UMP-CMP kinase, a nucleoside salvage pathway kinase. Like all NMP kinases, UMP-CMP kinase binds the phosphodonor, usually ATP, and the NMP at different binding sites. The reaction results from an in-line phosphotransfer from the donor to the acceptor. The probe for the donor site was displaced by the bisubstrate analogs of the Ap5X series (where X = U, dT, A, G), indicating the broad specificity of the acceptor site. Both CMP and dCMP were competitors for the acceptor site probe. To find antimetabolites for antivirus and anticancer therapies, we have developed a method of screening acyclic phosphonate analogs that is based on the affinity of the acceptor-binding site of the human UMP-CMP kinase. Several uracil vinylphosphonate derivatives had affinities for human UMP-CMP kinase similar to those of dUMP and dCMP and better than that of cidofovir, an acyclic nucleoside phosphonate with a broad spectrum of antiviral activities. The uracil derivatives were inhibitors rather than substrates of human UMP-CMP kinase. Also, the 5-halogen-substituted analogs inhibited the human TMP kinase less efficiently. The broad specificity of the enzyme acceptor-binding site is in agreement with a large substrate-binding pocket, as shown by the 2.1 A crystal structure.
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https://hal-pasteur.archives-ouvertes.fr/pasteur-00166127
Contributor : Sylvie Pochet <>
Submitted on : Wednesday, August 1, 2007 - 12:49:08 PM
Last modification on : Monday, January 13, 2020 - 5:08:07 PM

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Dimitri Topalis, Hiroki Kumamoto, Maria-Fernanda Amaya Velasco, Laurence Dugué, Ahmed Haouz, et al.. Nucleotide binding to human UMP-CMP kinase using fluorescent derivatives - a screening based on affinity for the UMP-CMP binding site.. FEBS Journal, Wiley, 2007, 274 (14), pp.3704-14. ⟨10.1111/j.1742-4658.2007.05902.x⟩. ⟨pasteur-00166127⟩

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