Cardiac ATP-sensitive K+ channel associates with the glycolytic enzyme complex
Cardiac ATP-sensitive K+ channel associates with the glycolytic enzyme complex
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DOI：
10.1096/fj.10-176669
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发表时间：
2011-07-01
期刊：
影响因子：
4.8
通讯作者：
Coetzee, William A.
中科院分区：
文献类型：
--
作者：
Hong, Miyoun;Kefaloyianni, Eirini;Coetzee, William A.
心脏ATP敏感性钾（K-ATP）通道是由高能核苷酸门控的，对细胞能量代谢的变化非常敏感。一个新兴的观点是，与K-ATP通道相关的蛋白质提供了额外的调节层。使用假定的磺酰脲受体（SUR）卷曲螺旋结构域作为诱饵，在2-杂交筛选对大鼠心脏cDNA文库，我们确定糖酵解酶（GAPDH和醛缩酶A）作为假定的相互作用蛋白。醛缩酶和SUR之间的相互作用，证实使用GST下拉试验和免疫共沉淀试验。从大鼠心肌膜的K-ATP通道免疫沉淀物的蛋白质质谱鉴定糖酵解作为最丰富的生物过程。免疫共沉淀试验证实了几种糖酵解酶在整个糖酵解途径中的相互作用。免疫细胞化学共定位许多这些酶与K-ATP通道亚单位在大鼠心肌细胞。在膜片钳实验中，醛缩酶和丙酮酸激酶的催化活性在功能上调节K-ATP通道，而D-葡萄糖没有影响。总的来说，我们的数据证明了糖酵解过程（特别是远端ATP生成步骤）与心脏K-ATP通道的密切物理联系和功能相互作用。洪，M.，Kefaloyianni，E.，鲍湖，加-地Malester，B.，Delaroche，D.，Neubert，T.一、Coetzee，W. A.心脏ATP敏感性K+通道与糖酵解酶复合物相关。FASEB J.25，2456-2467（2011）. www.fasebj.org
Being gated by high-energy nucleotides, cardiac ATP-sensitive potassium (K-ATP) channels are exquisitely sensitive to changes in cellular energy metabolism. An emerging view is that proteins associated with the K-ATP channel provide an additional layer of regulation. Using putative sulfonylurea receptor (SUR) coiled-coil domains as baits in a 2-hybrid screen against a rat cardiac cDNA library, we identified glycolytic enzymes (GAPDH and aldolase A) as putative interacting proteins. Interaction between aldolase and SUR was confirmed using GST pulldown assays and coimmunoprecipitation assays. Mass spectrometry of proteins from K-ATP channel immunoprecipitates of rat cardiac membranes identified glycolysis as the most enriched biological process. Coimmunoprecipitation assays confirmed interaction for several glycolytic enzymes throughout the glycolytic pathway. Immunocytochemistry colocalized many of these enzymes with K-ATP channel subunits in rat cardiac myocytes. The catalytic activities of aldolase and pyruvate kinase functionally modulate K-ATP channels in patch-clamp experiments, whereas D-glucose was without effect. Overall, our data demonstrate close physical association and functional interaction of the glycolytic process (particularly the distal ATP-generating steps) with cardiac K-ATP channels.- Hong, M., Kefaloyianni, E., Bao, L., Malester, B., Delaroche, D., Neubert, T. A., Coetzee, W. A. Cardiac ATP-sensitive K+ channel associates with the glycolytic enzyme complex. FASEB J. 25, 2456-2467 (2011). www.fasebj.org