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首页> 外文期刊>Journal of Molecular Biology >Impact of mutations affecting ND mitochondria-encoded subunits on the activity and assembly of complex I in Chlamydomonas. Implication for the structural organization of the enzyme.
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Impact of mutations affecting ND mitochondria-encoded subunits on the activity and assembly of complex I in Chlamydomonas. Implication for the structural organization of the enzyme.

机译：影响ND线粒体编码亚基的突变对衣藻中复合物I的活性和装配的影响。对酶的结构组织有影响。

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摘要

The mitochondrial rotenone-sensitive NADH:ubiquinone oxidoreductase (complex I) comprises more than 35 subunits, the majority of which are encoded by the nucleus. In Chlamydomonas reinhardtii, only five components (ND1, ND2, ND4, ND5 and ND6) are coded for by the mitochondrial genome. Here, we characterize two mitochondrial mutants (dum5 and dum17) showing strong reduction or inactivation of complex I activity: dum5 is a 1T deletion in the 3' UTR of nd5 whereas dum17 is a 1T deletion in the coding sequence of nd6. The impact of these mutations and of mutations affecting nd1, nd4 and nd4d5 genes on the assembly of complex I is investigated. After separation of the respiratory complexes by blue native (BN)-PAGE or sucrose gradient centrifugation, we demonstrate that the absence of intact ND1 or ND6 subunit prevents the assembly of the 850 kDa whole complex, whereas the loss of ND4 or ND4/ND5 leads to the formation of a subcomplex of 650 kDa present in reduced amount. The implications of our findings for the possible role of these ND subunits on the activity of complex I and for the structural organization of the membrane arm of the enzyme are discussed. In mitochondria from all the strains analyzed, we moreover detected a 160-210 kDa fragment comprising the hydrophilic 49 kDa and 76 kDa subunits of the complex I peripheral arm and showing NADH dehydrogenase activity.
机译：线粒体鱼藤酮敏感性NADH：泛醌氧化还原酶（复合体I）包含35个以上的亚基，其中大部分由细胞核编码。在莱茵衣藻中，线粒体基因组仅编码五个成分（ND1，ND2，ND4，ND5和ND6）。在这里，我们表征了两个线粒体突变体（dum5和dum17），它们显示了复杂I活性的强烈降低或失活：dum5是nd5的3'UTR中的1T缺失，而dum17是nd6的编码序列中的1T缺失。研究了这些突变以及影响nd1，nd4和nd4 / nd5基因的突变对复合物I装配的影响。通过蓝色天然（BN）-PAGE或蔗糖梯度离心分离呼吸道复合物后，我们证明了完整的ND1或ND6亚基的缺失阻止了850 kDa整个复合体的组装，而ND4或ND4 / ND5的缺失导致导致以减少的量存在的650 kDa的亚复合物的形成。讨论了我们的发现对这些ND亚基对复合物I的活性可能的作用以及对酶膜臂的结构组织的影响。此外，在所有分析菌株的线粒体中，我们检测到一个160-210 kDa的片段，该片段包含复合物I外围臂的亲水性49 kDa和76 kDa亚基，并显示出NADH脱氢酶活性。

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