Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.507205
Title: myo-Inositol 1,2,3-trisphosphate : iron chelation and conformational analysis
Author: Mansell, David James
Awarding Body: The University of Manchester
Current Institution: University of Manchester
Date of Award: 2009
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Abstract:
Mammalian cells contain a pool of iron not strongly bound to proteins. The cellular ligands of this biologically important 'chelatable', 'labile' or 'transit' iron are not known. Such ligands are expected to prevent iron from undergoing redox cycles that catalyse hydroxyl radical formation. Amongst small cellular molecules, myo-inositol phosphates containing the equatorial-axial-equatorial 1,2,3-trisphosphate grouping, are very effective at complexing Fe3+ in such a 'safe' manner ill vitro. myo-Inositol hexakisphosphate (InsP6, phytic acid), the most abundant inositol phosphate, was consequently recognised as a potential candidate for an intracellular Fe3+ ligand. However, thermodynamic data has recently excluded InsP6 as a potential intracellular Fe3+ complexing agent due to competition with the more abundant Mg2+ cations. It is not clear however, whether this same limitation also applies to myo-inositol 1,2,3- trisphosphate [Ins(l,2,3)P3], the simplest member of this group. Ins(1,2,3)P3 is a cellular constituent of unknown function, present in a variety of mammalian cells at concentrations of 1-10 JIM. The aim here was to synthesise Ins(I,2,3)P3 and study for the first time, its metal complexation behaviour under simulated cellular conditions. The stability constants of the comf-Iexes formed by Ins(I,2,3)P3 with biologically relevant metal cations (Na+, K+, Mg +, Ca2+, Cu2+, Fe2+ and Fe3+) are reported. These data indicate that Ins(1,2,3)P3 can be expected to complex to all available Fe3+, both in the cytosol/nucleus and acidic cellular compartments. The capability of Ins(1,2,3)P3 to inhibit iron redox cycling and associated production of free radicals is crucial, as redox-active iron should have been minimised by evolution. The binding conformation of Ins(1,2,3)P3 is predicted to be key to the ability of the 1,2,3-trisphosphate motif to bind iron in a 'safe' manner. Ironcoordinated Ins(1,2,3)P3 has been proposed to adopt the less stable penta-axial conformation, orientating the phosphate groups axial-equatorial-axiaL Structural examination of the Ins(I,2,3)P3-Fe3+ complex however, is severely restricted due to the paramagnetic properties of Fe3+ cations. In this work, the structure of the Ins(1,2,3)P-Fe3+ complex has been studied using a combination of approaches. The synthesis of 4,6-carbonate-myo-inositol 1,2,3,5-tetrakisphosphate as a model for the penta-axial conformation of the 1,2,3-trisphosphate motif is described. Subsequent iron binding studies with this molecule were comparable to that of Ins(1,2,3)P3. In contrast, the simple straight-chain analogue, trisphosphoglycerol, was unable to completely inhibit Fe3+-catalysed free radical formation. Furthermore, it was possible to monitor the conformation using a synthetic pyrene-based fluorescent probe, 4,6- bispyrenoyl-myo-inositol 1,2,3,5-tetrakisphosphate. Conformational ring-flip of the cyclohexane chair upon association with Fe3+ is accompanied by a dramatic change in fluorescence emission, due to 1C-1C stacking of the pyrene groups, promoting the formation of excimer fluorescence. High level quantum chemical calculations on the Ins(1,2,3)P3-Fe3+ complex revealed detailed structural information on the complex, predicting the involvement of inositol p/wsp/wester oxygens as well as terminal oxygens in the coordination of Fe3+. The findings here suggest Ins(1,2,3)P3 is the first viable proposal for an iron transit ligand. In addition, structural studies support the general principle that Fe3+ binds to the penta-axial conformation ofIns(l,2,3)P3.
Supervisor: Not available Sponsor: Not available
Qualification Name: Not available Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.507205  DOI: Not available
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