PHOSPHATIDYLINOSITOL SYNTHASE AND PHOSPHATIDYLINOSITOL INOSITOL EXCHANGE-REACTIONS IN TURKEY ERYTHROCYTE-MEMBRANES
PHOSPHATIDYLINOSITOL SYNTHASE AND PHOSPHATIDYLINOSITOL INOSITOL EXCHANGE-REACTIONS IN TURKEY ERYTHROCYTE-MEMBRANES
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DOI：
10.1042/bj2750187
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发表时间：
1991-04-01
影响因子：
4.1
通讯作者：
DOWNES, CP
中科院分区：
文献类型：
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作者：
MCPHEE, F;LOWE, G;DOWNES, CP
与人类红细胞不同，来自禽类的红细胞，如火鸡和小鸡，能迅速将肌-[H-3]肌醇纳入膜磷脂中。利用火鸡红细胞膜研究了[H-3]Ins标记磷脂酰肌醇的调控机制。在没有添加核苷酸的情况下，[H-3]肌醇掺入似乎是通过磷脂酰肌醇/肌醇交换进行的，肌醇的K(m)为0.01 mM。该反应依赖于二价阳离子，Mg2+或Mn2+，后一种金属离子更有效。[H-3] CMP加速了肌醇的掺入，特别是当Ins浓度大于交换反应的K(m)时。CMP依赖的PtdIns标记对肌醇的K(m)为0.3 mM，对CMP的K(m)为0.015 mM。该反应也需要二价阳离子：活性在0.5 mM- mn2 +时达到峰值，浓度越高，活性越低。在相对较高的浓度下，Mg2+比Mn2+更有效，在10 mM以上达到峰值活性。[H-3]肌醇的cmp依赖性结合似乎反映了由PtdIns合成酶催化的交换反应。通过证明[C-14]CMP形成[C-14]CMP-磷脂酸盐，获得了火鸡红细胞膜中PtdIns合成酶存在的明确证据。在未标记肌醇存在的情况下，放射性可以有效地从[C-14] cmp -磷酸酯中去除。在形态学简单的火鸡红细胞中检测PtdIns合成酶活性有助于阐明磷脂酰肌醇循环中这一重要组成部分的亚细胞分布。
Unlike human erythrocytes, those from avian species, such as turkeys and chicks, rapidly incorporate myo-[H-3]inositol into membrane phospholipids. The mechanisms regulating [H-3]Ins labelling of phosphatidylinositol have been investigated using turkey erythrocyte membranes. In the absence of added nucleotides, [H-3]inositol incorporation appears to proceed via phosphatidylinositol/inositol exchange, with a K(m) for inositol of 0.01 mM. The reaction was dependent upon divalent cations, either Mg2+ or Mn2+, with the latter metal ion being the more effective. [H-3]Inositol incorporation was accelerated by CMP, especially when the concentration of Ins was greater than the K(m) for the exchange reaction. CMP-dependent labelling of PtdIns had a K(m) for inositol of 0.3 mM and for CMP of 0.015 mM. Divalent cations were also required for this reaction: activity peaked at 0.5 mM-Mn2+ and declined at higher concentrations. At relatively high concentrations, Mg2+ was more effective than Mn2+, with peak activity being achieved above 10 mM. CMP-dependent incorporation of [H-3]inositol appears to reflect an exchange reaction catalysed by PtdIns synthase. Definitive evidence for the occurrence of PtdIns synthase in turkey erythrocyte membranes was obtained by demonstrating the formation of [C-14]CMP-phosphatidate from [C-14]CMP. The radioactivity could be efficiently chased from [C-14]CMP-phosphatidate in the presence of unlabelled inositol. The detection of PtdIns synthase activity in morphologically simple turkey erythrocytes should help to clarify the subcellular distribution of this important component of the phosphatidylinositol cycle.