Abstract
The inositol 1,4,5-trisphosphate (InsP3) receptor was purified from bovine cerebellum and reconstituted in liposomes composed of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) (1:1) successfully. No effect of Ca2+ concentration on [3H]-InsP3 binding to unreconstituted InsP3 receptor could be observed either at 4°C or at 25°C, whereas the effect of [Ca2+] on reconstituted InsP3 receptor depended on the temperature. The Ca2+ concentration outside the proteolipsome ([Ca2+]o) had no detectable effect on InsP3 binding to InsP3 receptor at 4°C. In contrast, with increase of [Ca2+]o from 0 to 100 nmol/L at 25°C, the InsP3 binding activity increased gradually. Then the InsP3 binding activity was decreased drastically at higher [Ca2+]o and inhibited entirely at 50 μmol/L [Ca2+]o. Conformational studies on intrinsic fluorescence of the reconstituted InsP3 receptor and its quenching by KI and HB indicated that the global conformation of reconstituted InsP3 receptor could not be affected by [Ca2+]o at 4°C. While at 25°C, the effects of 10 μmol/L [Ca2+]o on global, membrane and cytoplasmic conformation of the reconstituted InsP3 receptor were different significantly from that of 100 nmol/L [Ca2+]o.
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Hu, G., Huang, Y. & Yang, F. Temperature-dependence and conformational basis of inositol 1,4,5-trisphosphate receptor regulated by Ca2+ . Sci. China Ser. C.-Life Sci. 43, 225–231 (2000). https://doi.org/10.1007/BF02879280
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DOI: https://doi.org/10.1007/BF02879280