Chimeric proteins between UCP1 and UCP3: the middle third of UCP1 is necessary and sufficient for activation by fatty acids.
Chimeric proteins between UCP1 and UCP3: the middle third of UCP1 is necessary and sufficient for activation by fatty acids.
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UCP1 和 UCP3 之间的嵌合蛋白：UCP1 的中间三分之一对于脂肪酸的激活是必要且充分的。
DOI：
10.1006/bbrc.2000.3535
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发表时间：
2000
影响因子：
3.1
通讯作者：
Lowell,BB
中科院分区：
文献类型：
--
作者：
Hagen,T;Lowell,BB
解偶联蛋白1(UCP)1和解偶联蛋白3(UCP3)都是线粒体内膜蛋白，它们都介导质子泄漏，从而降低线粒体跨膜质子梯度。然而，UCP1和UCP3的生化调控不同。UCP1被游离脂肪酸激活，被嘌呤核苷酸抑制。利用酵母中的异源表达研究发现，UCP3既缺乏脂肪酸激活，也缺乏嘌呤核苷酸抑制。为了评估哪些结构域负责由游离脂肪酸和嘌呤核苷酸调节UCP1，以及UCP3中没有这种调节，产生了嵌合蛋白。由于解偶联蛋白像线粒体载体家族的所有成员一样，具有三方结构，由大约100个残基的三个重复结构域组成，UCP1和UCP3在这三个重复结构域之间进行了互换。在酵母线粒体中异源表达后，研究了游离脂肪酸和嘌呤核苷酸对6种不同嵌合蛋白的调节作用。本研究表明，由于UCP1的第二重复序列被UCP3的等效重复序列所替代，因此UCP1被游离脂肪酸激活是由第二重复结构域介导的。相反，用UCP1的相应重复结构域取代UCP3的第二个重复结构域会导致类似于野生型UCP1的脂肪酸激活。UCP3缺乏游离脂肪酸激活并不是因为在UCP1的第二重复结构域中没有组氨酸对H145和H147。此外，关于嘌呤核苷酸抑制的研究结果与UCP1的C末端重复结构域在介导嘌呤核苷酸抑制中的显著作用是一致的。
Uncoupling protein (UCP) 1 and UCP3 are mitochondrial inner membrane proteins which both mediate proton leak and thus decrease the mitochondrial transmembrane proton gradient. However, UCP1 and UCP3 differ in their biochemical regulation. UCP1 is activated by free fatty acids and inhibited by purine nucleotides. Using heterologous expression studies in yeast, UCP3 was found to lack both fatty acid activation and purine nucleotide inhibition. To assess which domains are responsible for the regulation of UCP1 by free fatty acids and by purine nucleotides and the absence of such regulation in UCP3, chimeric proteins were generated. Given that uncoupling proteins, like all members of the mitochondrial carrier family, possess a tripartite structure and consist of three repeated domains of approximately 100 residues, swaps in the three repeated domains were made between UCP1 and UCP3. Regulation of the resulting six different chimeric proteins by free fatty acids and purine nucleotides was studied after heterologous expression in yeast mitochondria. In this study, it is shown that activation of UCP1 by free fatty acids is mediated by the second repeated domain, since substitution of the second repeat of UCP1 by the equivalent repeat of UCP3 abolishes fatty acid activation. In contrast, replacing the second repeat of UCP3 by the corresponding repeated domain of UCP1 results in fatty acid activation similar to wild type UCP1. The lack of free fatty acid activation of UCP3 is not due to the absence of the histidine pair H145 and H147 found in the second repeated domain of UCP1. Furthermore, the findings with respect to purine nucleotide inhibition are consistent with a significant role of the C-terminal repeated domain of UCP1 in mediating purine nucleotide inhibition.