3-Position modification of myo-inositol 1,4,5-trisphosphate: consequences for intracellular Ca²⁺ mobilisation and enzyme recognition

The ability of the novel D-myo-inositol 1,4,5-trisphosphate (Ins(1,4,5)P₃) analogues, L-chiro-inositol 2,3,5-trisphosphate (L-ch-Ins(2,3,5)P₃) and D-3-deoxy-3-fluoro-myo-inositol 1,4,5-trisphophate (3F-Ins(1,4,5)P₃), to bind to the Ins(1,4,5)P₃ receptor, mobilise intracellular Ca²⁺ stores and interact with metabolic enzymes has been investigated. L-ch-Ins(2,3,5)P₃ and 3F-Ins(1,4,5)P₃ were bound by the Ins(1,4,5)P₃ receptor from bovine adrenal cortex with relatively high affinity (K_i values 60.4 and 8.0 nM respectively) but with lower affinity than Ins(1,4,5)P₃ (K_D = 5.9 nM). Both analogues were apparent full agonists at the Ca²⁺ mobilising receptor in SH-SY5Y cells, but were less potent than Ins(1,4,5)P₃ (EC₅₀ L-ch-Ins(2,3,5)P₃ = 1.4 μM,F-Ins(1,4,5)P₃ = 0.37 μM and Ins(1,4,5)P₃ = 0.12 μM). L-ch-Ins(2,3,5)P₃ and 3F-Ins(1,4,5)P₃ were resistant to Ins(1,4,5)P₃ 3-kinase, and were potent inhibitors of the enzyme (K_i values 7.1 and 8.6 μM respectively). 3F-Ins(1,4,5)P₃ was hydrolysed by Ins(1,4,5)P₃ 5-phosphatase at a similar rate to Ins(1,4,5)P₃, but inhibited dephosphorylation of [³H]Ins(1,4,5)P₃ with high potency (apparent K_i = 3.9 μM) L-ch-Ins(2,3,5)P₃ was also recognised by the enzyme with high affinity (K_i = 7.7 μM) but was resistant to hydrolysis. These results suggest that the environment around C-3 is of major importance for recognition not only by Ins(1,4,5)P₃ 3-kinase but also by Ins(1,4,5)P₃ 5-phosphatase.