Flexibility in Anaerobic Metabolism as Revealed in a Mutant of Chlamydomonas reinhardtii Lacking Hydrogenase Activity
Flexibility in Anaerobic Metabolism as Revealed in a Mutant of Chlamydomonas reinhardtii Lacking Hydrogenase Activity
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DOI：
10.1074/jbc.m803917200
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发表时间：
2009-03-13
影响因子：
4.8
通讯作者：
Posewitz, Matthew C.
中科院分区：
文献类型：
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作者：
Dubini, Alexandra;Mus, Florence;Posewitz, Matthew C.
绿色衣藻有一个发酵途径网络，当细胞适应缺氧时，这个网络变得活跃。氢化酶活性是这种代谢的重要组成部分，我们比较了野生型C. reinhardtii细胞和编码[FeFe]氢化酶成熟蛋白的HYDEF基因的无效突变株（hydEF-1突变体）。该突变体没有氢化酶活性，并且在黑暗厌氧代谢期间相对于亲本菌株表现出琥珀酸积累增加和CO2产生减少。在缺乏氢化酶活性的情况下，琥珀酸积累增加表明细胞激活丙酮酸代谢的替代途径，这有助于NAD（P）H再氧化，并在缺乏O-2的情况下继续糖酵解和发酵。发酵性琥珀酸生产可能通过苹果酸的形成进行，并且在细胞从有氧条件转移到缺氧条件后，相对于亲代菌株，在突变体中观察到编码两种苹果酸形成酶（丙酮酸羧化酶和苹果酸酶）的mRNA丰度增加。虽然C.莱茵衣藻具有编码丙酮酸羧化酶的单个基因，它具有编码推定的苹果酸酶的六个基因。只有一个苹果酸酶基因，MME 4，表现出显着增加的表达（mRNA丰度）的hydEF-1突变体在厌氧。此外，有显着增加的转录物编码的脱氢酶和延胡索酸还原酶，酶purified需要将苹果酸转化为琥珀酸。这些结果说明了C. reinhardtii的，并有助于在这种和潜在的其他藻类的厌氧代谢的知情模型的发展。
The green alga Chlamydomonas reinhardtii has a network of fermentation pathways that become active when cells acclimate to anoxia. Hydrogenase activity is an important component of this metabolism, and we have compared metabolic and regulatory responses that accompany anaerobiosis in wild-type C. reinhardtii cells and a null mutant strain for the HYDEF gene (hydEF-1 mutant), which encodes an [FeFe] hydrogenase maturation protein. This mutant has no hydrogenase activity and exhibits elevated accumulation of succinate and diminished production of CO2 relative to the parental strain during dark, anaerobic metabolism. In the absence of hydrogenase activity, increased succinate accumulation suggests that the cells activate alternative pathways for pyruvate metabolism, which contribute to NAD(P)H reoxidation, and continued glycolysis and fermentation in the absence of O-2. Fermentative succinate production potentially proceeds via the formation of malate, and increases in the abundance of mRNAs encoding two malate-forming enzymes, pyruvate carboxylase and malic enzyme, are observed in the mutant relative to the parental strain following transfer of cells from oxic to anoxic conditions. Although C. reinhardtii has a single gene encoding pyruvate carboxylase, it has six genes encoding putative malic enzymes. Only one of the malic enzyme genes, MME4, shows a dramatic increase in expression (mRNA abundance) in the hydEF-1 mutant during anaerobiosis. Furthermore, there are marked increases in transcripts encoding fumarase and fumarate reductase, enzymes putatively required to convert malate to succinate. These results illustrate the marked metabolic flexibility of C. reinhardtii and contribute to the development of an informed model of anaerobic metabolism in this and potentially other algae.