pInsiP/isub3/sub binding to type-1, but not type-3, InsiP/isub3/sub receptors is inhibited by calmodulin in a Casup2+/sup-independent fashion [Cardy and Taylor (1998) Biochem. J. b334/b, 447-455], and Casup2+/sup mobilization by type-1 InsiP/isub3/sub receptors of cerebellum is inhibited by calmodulin [Patel, Morris, Adkins, O9Beirne and Taylor (1997) Proc. Natl. Acad. Sci. U.S.A. b94/b, 11627-11632]. Using cell types expressing predominantly type-1, -2 or -3 InsiP/isub3/sub receptors, we show that InsiP/isub3/sub-evoked Casup2+/sup mobilization from each is similarly inhibited by calmodulin. In SH-SY5Y cells, which express largely type-1 receptors, calmodulin (ICsub50/sub ≈ 15 iμ/iM) inhibited InsiP/isub3/sub-evoked Casup2+/sup release only in the presence of Casup2+/sup. The inhibition was unaffected by calcineurin inhibitors. The effect of calmodulin did not result from enhanced metabolism of InsiP/isub3/sub because calmodulin also decreased the sensitivity of the Casup2+/sup stores to adenophostin A, a non-metabolizable InsiP/isub3/sub-receptor agonist. Protein kinase A-catalysed phosphorylation of type-1 InsiP/isub3/sub receptors was unaffected by Casup2+/sup-calmodulin. Using a scintillation proximity assay to measure sup125/supI-calmodulin binding to glutathione S-transferase-fusion proteins, we identified two regions of the type-1 InsiP/isub3/sub receptor (cyt1, residues -6 to 159; and cyt11, residues 1499-1649) that bound sup125/supI-calmodulin. The higher-affinity site (cyt11) was also photoaffinity labelled with iN/i-hydroxysuccinimidyl-4-azidobenzoate (HSAB)-calmodulin. We speculate that Casup2+/sup-independent binding of calmodulin to a site within the first 159 residues of the type-1 InsiP/isub3/sub receptor inhibits InsiP/isub3/sub binding and may thereby regulate the kinetics of Casup2+/sup release. Casup2+/sup-dependent inhibition of Casup2+/sup release by calmodulin is mediated by a different site: it may reside on an accessory protein that associates with all three receptor subtypes, or Casup2+/sup-calmodulin binding to a site lying between residues 1499 and 1649 of the type-1 receptor may inhibit Casup2+/sup release from any tetrameric receptor that includes a type-1 subunit./p
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机译：> Ins P 3 与1型而非3型Ins P 3 受体结合钙调蛋白以Ca 2+无关的方式抑制钙-磷[Cardy and Taylor（1998）Biochem。 J. 334 ，447-455]和1型Ins P 3 受体动员Ca 2 + 小脑的抑制被钙调蛋白抑制[Patel，Morris，Adkins，O9Beirne和Taylor（1997）Proc.Natl.Acad.Sci.USA 90：5873-5877。 Natl。学院科学U.S.A. 94 ，11627-11632]。使用主要表达类型1，-2或-3 Ins P 3 受体的细胞类型，我们显示Ins P 3 <钙调蛋白同样抑制了每个/ sub诱发的Ca 2 + 动员。在主要表达1型受体的SH-SY5Y细胞中，钙调蛋白（IC 50 ≈15 μ M）抑制Ins P 3 诱发的Ca 2 + 仅在Ca 2 + 存在时释放。该抑制不受钙调神经磷酸酶抑制剂的影响。钙调蛋白的作用不是由Ins P 3 的新陈代谢引起的，这是因为钙调蛋白还降低了Ca 2 + 储库对腺苷A的敏感性，一种不可代谢的Ins P 3 -受体激动剂。蛋白激酶A催化1型Ins P 3 受体的磷酸化不受Ca 2 + -钙调蛋白的影响。使用闪烁邻近测定法测量 125 I-钙调蛋白与谷胱甘肽S-转移酶融合蛋白的结合，我们确定了1型Ins P 3的两个区域结合 125 I-钙调蛋白的受体（cyt1，残基-6至159; cyt11，残基1499-1649）。较高亲和力的位点（cyt11）也用N-羟基琥珀酰亚胺基-4-叠氮苯甲酸酯（HSAB）-钙调蛋白进行光亲和标记。我们推测钙调蛋白与Ca 2 + 的独立结合与1型Ins P 3 受体的前159个残基内的位点具有抑制作用Ins P 3 的结合并可能调节Ca 2 + 释放的动力学。钙调蛋白对Ca 2 + 依赖性的Ca 2 + 释放的抑制作用是由不同的位点介导的：它可能驻留在与所有三种受体亚型相关的辅助蛋白上，或Ca 2 + 钙调蛋白结合到1型受体残基1499和1649之间的位点可能会抑制Ca 2 + 从任何包括以下分子的四聚体受体中释放： 1个亚基。
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