title: Identification and Preliminary Characterization of a Ca2+- Dependent High-Affinity Binding Site for Inositol-1,4,5-Trisphosphate from Chenopodium rubrum.
abstract: Using a radioligand-binding assay we have identified a Ca2+- dependent high-affinity D-myo-inositol-1,4,5-trisphosphate (InsP3) binding site in a membrane vesicle preparation from Chenopodium rubrum. Millimolar concentrations of Ca2+ were required to observe specific binding of [3H]InsP3. A stable equilibrium between bound and free ligand was established within 5 min and bound [3H]InsP3 could be completely displaced by InsP3 in a time- and concentration-dependent manner. Displacement assays indicated a single class of binding sites with an estimated dissociation constant of 142 [plus or minus] 17 nM. Other inositol phosphates bound to the receptor with much lower affinity. The glycosaminoglycan heparin was an effective competitor for the binding site (inhibitor concentration for 50% displacement = 534 nM). ATP at higher, although physiologically relevant, concentrations (inhibitor concentration for 50% displacement = 241 [mu]M) also displaced [3H]InsP3 from the receptor. Recent studies in animals have highlighted the importance of Ca2+ regulation of InsP3-induced Ca2+ release. The potential for the operation of similar regulatory mechanisms in plants is discussed.
authors.affiliation: Department of Applied Biology, University of Central Lancashire, Preston, PR1 2HE, United Kingdom (C.H.S., C.E.R., P.J.L).
authors.forename: C. H.
authors.initials: CH
authors.lastname: Scanlon
authors.forename: J.
authors.initials: J
authors.lastname: Martinec
authors.forename: I.
authors.initials: I
authors.lastname: Machackova
authors.forename: C. E.
authors.initials: CE
authors.lastname: Rolph
authors.forename: P. J.
authors.initials: PJ
authors.lastname: Lumsden