Description<unk>  is a "suicide inhibitor," irreversibly binding to ornithine decarboxylase (ODC) and preventing the natural substrate ornithine from accessing the active site (Figure 1). Within the active site of ODC,<unk>  undergoes decarboxylation with the aid of cofactor pyridoxal 5'-phosphate (PLP). Because of its additional difluoromethyl group in comparison to ornithine,<unk>  is able to bind to a neighboring Cys-360 residue, permanently remaining fixated within the active site. During the reaction,<unk> 's decarboxylation mechanism is analogous to that of ornithine in the active site, where transamination occurs with PLP followed by decarboxylation. During the event of decarboxylation, the fluoride atoms attached to the additional methyl group pull the resulting negative charge from the release of carbon dioxide, causing a fluoride ion to be released. In the natural substrate of ODC, the ring of PLP accepts the electrons that result from the release of CO2. The remaining fluoride atom that resides attached to the additional methyl group creates an electrophilic carbon that is attacked by the nearby thiol group of Cys-360, allowing<unk>  to remain permanently attached to the enzyme following the release of the second fluoride atom and transimination. Evidence The reaction mechanism of Trypanosoma bruceis ODC with ornithine was characterized by UV-VIS spectroscopy in order to identify unique intermediates that occurred during the reaction. The specific method of multiwavelength stopped-flow spectroscopy utilized monochromatic light and fluorescence to identify five specific intermediates due to changes in absorbance measurements. The steady-state turnover number, kcat, of ODC was calculated to be 0.5 s−1 at 4 °C. From this characterization, the rate-limiting step was determined to be the release of the product putrescine from ODC's reaction with ornithine. In studying the hypothetical reaction mechanism for<unk> , information collected from radioactive peptide and<unk>  mapping, high pressure liquid chromatography, and gas phase peptide sequencing suggested that Lys-69 and Cys-360 are covalently bound to<unk>  in T. brucei ODC's active site. Utilizing fast-atom bombardment mass spectrometry (FAB-MS), the structural conformation of<unk>  following its interaction with ODC was determined to be (S)-((2-(1-pyrroline-methyl) cysteine, a cyclic imine adduct. Presence of this particular product was supported by the possibility to further reduce the end product to (S)-((2-pyrrole) methyl) cysteine in the presence of NaBH4 and oxidize the end product to (S)-((2-pyrrolidine) methyl) cysteine (Figure 2). Active site<unk> 's suicide inhibition of ODC physically blocks the natural substrate ornithine from accessing the active site of the enzyme (Figure 3). There are two distinct active sites formed by the homodimerization of ornithine decarboxylase. The size of the opening to the active site is approximately 13.6 Å. When these openings to the active site are blocked, there are no other ways through which ornithine can enter the active site. During the intermediate stage of<unk>  with PLP, its position near Cys-360 allows an interaction to occur. As the phosphate of PLP is stabilized by Arg 277 and a Gly-rich loop (235-237), the difluoromethyl group of<unk>  is able to interact and remain fixated to both Cys-360 and PLP prior to transimination. As shown in the figure, the pyrroline ring interferes with ornithine's entry (Figure 4).<unk>  will remain permanently bound in this position to Cys-360. As ODC has two active sites, two<unk>  molecules are required to completely inhibit ODC from ornithine decarboxylation. File:Ornithine and<unk>  Comparison Structures.Figure 1(A) 3D structure of L-Ornithine (B) 3D structure of<unk> . This molecule is similar to the structure of L-Ornithine, but its alpha-difluoromethyl group allows interaction with Cys-360 in the active site File:Eflornithine Reaction Mechansim.Eflornithine ODC reaction mechanism File:DFMO inhibitor product evidence.Figure 2 Experimental Evidence for<unk>  End Product File:Eflornithine in the Active Site.Figure 3 Active Site of ODC Formed by Homodimerization (Green and White Surface Structures) (A) Ornithine in the Active Site of ODC, Cys-360 highlighted in yellow (B) Product of<unk>  Decarboxylation bound to Cys 360 (highlighted in yellow). The pyrroline ring blocks ornithine from entering the active site
Eflornithine
Dimethyl methylphosphonate
Sodium azide
Mesocarb
Isocarboxazid
Metamizole
Fenvalerate
Fursultiamine