Complexes between porin, hexokinase, mitochondrial creatine kinase and adenylate translocator display properties of the permeability transition pore.: Implication for regulation of permeability transition by the kinases
Complexes between porin, hexokinase, mitochondrial creatine kinase and adenylate translocator display properties of the permeability transition pore.: Implication for regulation of permeability transition by the kinases
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DOI：
10.1016/s0005-2736(97)00175-2
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发表时间：
1998-01-05
影响因子：
3.4
通讯作者：
Brdiczka, D
中科院分区：
文献类型：
--
作者：
Beutner, G;Rück, A;Brdiczka, D
最近发现己糖激酶、外膜孔蛋白和腺苷酸转位酶（ANT）之间形成的复合物在一个重组系统中具有线粒体通透性转换孔的特性。该复合物是用0.5%的Triton X - 100从大鼠脑膜中提取出来的，并通过阴离子交换色谱法进行分离。其分子量约为400 kDa，表明是己糖激酶的四聚体（单体为100 kDa）。通过同样的方法，一种孔蛋白、肌酸激酶八聚体、ANT复合物被分离出来并在脂质体中重组。含有重组复合物的囊泡都保留了ATP，这些ATP可被两种激酶中的任何一种用于磷酸化外部的肌酸或葡萄糖。苍术苷抑制了这种活性，表明ANT参与了这一过程并且在功能上得到了重组[1]。仅从含有己糖激酶复合物的脂质体中，内部的苹果酸或ATP会以一种对N - 甲基缬氨酸 - 4 - 环孢菌素敏感的方式因添加Ca²⁺而释放，这表明己糖激酶 - 孔蛋白 - ANT复合物可能包含通透性转换孔（PTP）。这种类似PTP结构的Ca²⁺依赖性开放被ADP（表观I - 50，8 μM）和ATP（表观I - 50，84 μM）所抑制。葡萄糖也抑制了类似PTP的活性，而6 - 磷酸葡萄糖则消除了这种效应。尽管孔蛋白和ANT在含有肌酸激酶八聚体复合物的囊泡中具有功能活性，但Ca²⁺并没有诱导内部底物的释放。然而，在肌酸激酶八聚体解离后，该复合物表现出类似PTP的特性，并且在添加Ca²⁺时囊泡会释放内部代谢物。后一过程也被N - 甲基缬氨酸 - 4 - 环孢菌素所抑制。在复合物分离过程中对肽基 - 脯氨酰 - 顺反异构酶（代表亲环素）的活性进行了监测。亲环素D与己糖激酶复合物共同纯化，而在肌酸激酶复合物中不存在。N - 甲基缬氨酸 - 4 - 环孢菌素对肌酸激酶复合物的抑制作用可以通过与似乎支持八聚体形成的肌酸激酶二聚体的直接相互作用来解释。（C）1998爱思唯尔科学出版社
请注意，其中一些专业术语可能需要根据具体学科背景进一步准确理解和调整，比如“hexokinase”（己糖激酶）、“porin”（孔蛋白）、“adenylate translocator”（腺苷酸转位酶）、“creatine kinase”（肌酸激酶）等。
Complexes between hexokinase, outer membrane porin, and the adenylate translocator (ANT) were recently found to establish properties of the mitochondrial permeability transition pore in a reconstituted system. The complex was extracted by 0.5% Triton X-100 from rat brain membranes and separated by anion exchanger chromatography. The molecular weight was approximately 400 kDa suggesting tetramers of hexokinase (monomer 100 kDa). By the same method a porin, creatine kinase octamer, ANT complex was isolated and reconstituted in liposomes. Vesicles containing the reconstituted complexes both retained ATP that could be used by either kinase to phosphorylate external creatine or glucose. Atractyloside inhibited this activity indicating that the ANT was involved in this process and was functionally reconstituted [1]. Exclusively from the hexokinase complex containing liposome internal malate or ATP was released by addition of Ca2+ in a N-methylVal-4-cyclosporin sensitive way, suggesting that the hexokinase porin ANT complex might include the permeability transition pore (PTP). The Ca2+ dependent opening of the PTP-like structure was inhibited by ADP (apparent I-50, 8 mu M) and ATP (apparent I-50, 84 mu M). Also glucose inhibited the PTP-like activity, while glucose-6-phosphate abolished this effect. Although porin and ANT were functionally active in vesicles containing the creatine kinase octamer complex, Ca2+ did not induce a release of internal substrates. However, after dissociation of the creatine kinase octamer, the complex exhibited PTP-like properties and the vesicles liberated internal metabolites upon addition of Ca2+. The latter process was also inhibited by N-methylVal-4-cyclosporin. The activity of peptidyl-prolyl-cis-trans-isomerase (representing cyclophilin) was followed during complex isolation. Cyp D was co-purified with the hexokinase complex, while it was absent in the creatine kinase complex. The inhibitory effect of N-methylVal-4-cyclosporin on the creatine kinase complex may be explained by direct interaction with the creatine kinase dimer that appeared to support octamer formation. (C) 1998 Elsevier Science B.V.