粟酒裂殖酵母中Atp11在线粒体中功能的研究

Study on the Function of Atp11 in Schizosaccharomyces pombe

ATP是由ATP合酶复合体催化合成的,在线粒体呼吸过程中起着重要作用。旨在研究粟酒裂殖酵母中Atp11(SPAC3A12.12)在线粒体中的功能。利用同源重组的方法构建Δatp11突变体,观察Δatp11缺失菌在以甘油为唯一碳源的非发酵培养基上的表型;通过生物信息学分析发现Atp11的N端含有一段由24个氨基酸残基组成的线粒体定位序列(MTS),为确定Atp11在细胞内的定位,在Atp11的C端添加一个GFP标签,观察绿色荧光在细胞内的位置;运用Western blotting检测atp11的缺失对线粒体基因组编码蛋白稳态性的影响。研究结果表明,Δatp11突变体在非发酵培养基上表现出生长缺陷,是线粒体呼吸缺陷菌;GFP绿色荧光定位实验结果证实了Atp11定位于线粒体中;Western blotting检测结果显示atp11的缺失导致Cob1、Cox1、Cox2、Cox3和Atp6蛋白的表达量显著降低。综上所述,粟酒裂殖酵母Atp11定位于线粒体且是线粒体呼吸链正常发挥功能所必须的。

ATP is synthesized by the catalase of ATP synthase complex and plays an important role in mitochondrial respiration.This paper aims to reveal the main function of Atp11 protein in Schizosaccharomyces pombe.First,we constructed the mutantΔatp11 by homologous recombination and observed the growth phenotype ofΔatp1-deleted strain in non-fermentative medium with glycerol as the sole carbon source.Second,we utilized bioinformatics analysis and discovered that Atp11 contained a 24-amino-acid mitochondrial localization sequence(MTS)at the N-terminus.To further confirm the location of Atp11 in the cell,a GFP label was added to the C-terminus of Atp11 to observe the position of green fluorescence in cells.Third,we used Western blotting to detect the effect of atp11 deletion on the stabilities of mitochondrial genomeencoded proteins.The results showed thatΔatp11 strain presented growth defect on non-fermentation medium and was a strain with faulted mitochondrial respiration.GFP experiment confirmed that Atp11 was localized in mitochondria.Western blotting experiment revealed that the deletion of atp11 resulted in the significant decrease in the protein level of Cob1,Cox1,Cox2,Cox3 and Atp6.In conclusion,Atp11 is localized in the mitochondria and essential for the function of mitochondrial respiration chain in S.pombe.