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题名	蓝藻碳代谢途径两个关键酶FBA和FBPase的进化研究
作者	汪德勇
学位类别	博士
答辩日期	2013-11
授予单位	中国科学院研究生院
授予地点	北京
导师	文建凡
关键词	蓝藻 果糖-1 碳代谢 6-二磷酸酶（FBPase） 果糖-1 进化 6-二磷酸缩醛酶（FBA）
其他题名	Studies on the two carbon metabolic enzymes of cyanobacteria——FBA and FBPase
学位专业	细胞生物学
中文摘要	光合作用是地球上最重要的生物化学反应。通过光合作用，以高等植物为代表的大量光合生物将太阳能转化为化学能，同时将空气中的CO2固定为糖类，为包括人在内的其他大量生物提供最初的物质和能量来源。放氧型光合作用起源于原核生物蓝藻，后者通过内共生作用，逐渐演化成今天真核藻类和植物细胞中专门进行光合作用的细胞器——叶绿体。而蓝藻仍然是现存唯一可以进行放氧光合作用的原核生物。因此，对参与蓝藻光合作用的相关基因的进化进行研究，有助于加深我们对光合作用进化规律的认识。本研究选取蓝藻光合作用暗反应——卡尔文循环中的两个关键酶果糖-1，6-二磷酸缩醛酶（FBA）和果糖-1，6-二磷酸酶（FBPase），对与它们的进化问题进行了比较深入的研究，得到如下结果和结论： 1）在对蓝藻FBA的研究中我们首先发现：与FBA在整个生物界的大致分布趋势即真核生物一般具有FBA I而原核生物一般具有FBA II相符，绝大部分蓝藻的基因组中都具有一个FBA II基因。然而，不少蓝藻的基因组中不仅具有FBA II基因，还具有FBA I基因，有两种蓝藻的基因组中甚至还具有不只一个FBA I基因。更有意思的是，在最近分化出来的四株原绿球藻的基因组中没有FBA II基因，却具有一个FBA I基因。从FBA在蓝藻中的这一分布模式我们初步推测：FBA II是蓝藻的原初的古老型FBA酶，而FBA I是一些蓝藻在后来进化过程中次生性获得的，四株原绿球藻在进化过程中甚至丢失了原有的FBA II基因而只保留了后来获得的FBA I基因。为检验这一推测，我们用包括蓝藻和最近鉴定到的噬藻体FBA I在内的很多物种的FBA I蛋白序列进行系统发生分析，结果不仅确定了蓝藻FBA I基因是后来获得的，更进一步发现其具有多个独立的起源。更有意思的是发现，最近鉴定到的噬藻体来源的FBA I蛋白序列与海洋属聚球藻和原绿球藻的聚在一起，两种gamma蛋白菌的FBA I蛋白序列构成它们的直接外群。这说明这些海洋属聚球藻和原绿球藻的FBA I基因是通过噬藻体为中介由gamma蛋白菌转移来的。这一发现也推翻了前人认为的这两类蓝藻的FBA I基因是直接从红藻转移而来的结论。更进一步对FBA I基因在蓝藻基因组中所处位置的上下文进行了比较分析，结果不仅支持上述蓝藻FBA I基因为多次独立起源的结论，而且还表明蓝藻获得FBA I基因可能是通过更多次的水平基因转移事件完成的。这些研究首次对蓝藻FBA I基因的起源进行了全面的调查，确定了蓝藻FBA I基因具有多个独立起源，其获得FBA I基因可能是通过更多次的水平基因转移事件完成的，噬藻体在海洋属聚球藻和原绿球藻获得FBA I基因的过程中起着十分重要的中介作用。 2）在对蓝藻FBPase的研究中，与FBA相似，蓝藻中也存在FBPase I和FBPase II两种没有同源性的FBPase。其中FBPase I与高等植物的FBPase（分为细胞质型FBPase和叶绿体型FBPase）同源，是一个单功能酶；而FBPase II是一个只在原核生物中存在的酶，在蓝藻中其还是一个双功能酶，同时具有FBPase和SBPase的酶活性。那么蓝藻中FBPase的这种分布格局是如何进化形成的呢？我们对蓝藻中这两种类型的FBPase的分布作了全面的调查，发现FBPase II在蓝藻中普遍存在，几乎每一个蓝藻的基因组中都有一个FBPase II基因；而FBPase I只在部分蓝藻基因组中存在。更进一步对蓝藻FBPase II氨基酸序列的序列特征进行了仔细分析，发现有很多蓝藻的FBPase II的氨基酸序列中都具有一个保守的插入片段，而有些蓝藻的FBPase II却没有这种插入片段。在有插入片段的蓝藻FBPase II中，绝大部分（两种除外）都含有一对保守的半胱氨酸。此外，FBPase II氨基酸序列中的插入片段和保守半胱氨酸对的有无还与基因组中编码FBPase I基因的有无之间有很好的互补对应关系。联想到这与高等植物中细胞质型FBPase和叶绿体型FBPase之间的功能互补关系的相似性，我们推测蓝藻FBPase II氨基酸序列中的插入片段和其中的保守半胱氨酸对可能也像高等植物叶绿体型FBPase一样，参与对其酶活性的光调节。为此，我们对几种代表性蓝藻的FBPase进行了体外表达，并研究其酶活性调节性质。初步结果表明：即具有插入片段又具有保守半胱氨酸的蓝藻FBPase II的酶活性可能是受到光调节的。这说明在一些现存最古老的蓝藻中FBPase II可能已经具有了完整的酶活性光调节系统，但是后来进化的蓝藻又为何丢失了这一调节系统并丢失了FBPase I，还有待于将来进一步研究。
英文摘要	The photosynthesis is the most important biochemical reaction on the world, through which the higher land plants and many other organisms transfer the solar light energy into the biochemical energy, accompanied with transferring the carbon dioxide into the carbohydrates. These carbohydrates as well as the energy contained in them constitute the foundation of animal survival. The oxygenic photosynthesis arised from the cyanobacteria, thereafter it transferred to the higher land plants and evolved into the chloroplast through the endosymbiosis. On the other hand, the cyanobacteria is still the only prokaryotes which possess the ability of oxygenic photosynthesis. Thus, the research on the evolution of cyanobacterial photosynthetic genes will deepen our understanding about the evolutionary mechanism of photosynthesis. In this study, we investigated two important carbon metabolic enzymes of cyanobacteria, that is, the fructose-1, 6-bisphosphate aldolase (FBA) and fructose-1, 6-bisphosphatase (FBPase). We got many innovative and meaningful results and conclusions which were presented below: 1) In the research of cyanobacterial FBA, we found that most cyanobacteria possess one FBA II gene in each genome which was consistent with the universal FBA distribution pattern in that the prokaryotes typically contain FBA II gene and eukaryotes typically contain FBA I gene. However, we also found that a small number of cyanobacteria possess FBA I genes in addition to the FBA II genes, and two cyanobacterial species possess more than one FBA I genes in their genomes. The most interesting was that we found four recently-evolving Prochlorococcus strains possess only the FBA I gene without the FBA II gene. Thus we speculated that the FBA II was the ancestral FBA enzymes of cyanobacteria and a small number of cyanobacteria acquired FBA I gene in their own evolutionary history. Moreover, four recently-evolving Prochlorococcus strains lost the FBA II genes and only retained the newly-acquired FBA I genes. We further reconstructed the FBA I phylogenetics using the FBA I protein sequences derived from all the cyanobacteria investigated in this study, the recently identified viral-derived FBA I sequences and many other organisms. The results of this part not only supported that the FBA I genes was transferred into some cyanobacterial species/strains in their own evolutionary history, but also indicated that there were multiple independent origins of cyanobacterial FBA I genes. What’s more interesting was that we found the recently identified viral-derived FBA I sequences clustered together with FBA I clade containing both marine Synechococcus and Prochlorococcus. The FBA I of two gamma proteobacteria constituted the immediate outgroup of these FBA I sequences. These findings suggested that the FBA I of marine Synechococcus and Prochlorococcus were acquired from gamma proteobacteria via virus intermediate which was contrast with previous suggestion that the FBA I of marine Synechococcus and Prochlorococcus were transferred from plastid-targeted red algae genes. At last, we investigated the genomic context of these newly acquired cyanobacterial FBA I genes. The results indicated that there may be more times of independent transfer events of FBA I genes to cyanobacteria. Overall, we suggested that the cyanobacterial FBA I genes have multiple independent origins with more times of independent transfer events, and the FBA I genes of marine Synechococcus and Prochlorococcus transferred from gamma proteobacteria instead of red algae via virus intermediate. 2) In the research of cyanobacterial FBPase, first we found that cyanobacteria possess two non-homologous FBPase: FBPase I and FBPase II, in which the FBPase I is homologous to the higher land plants FBPase (cytosolic FBPase and chloroplast FBPase) and FBPase II is specific to prokaryotes. The FBPase I is a unifunctional enzyme and FBPase II is a bifunctional enzyme possessing FBPase/SBPase activities.
语种	中文
内容类型	学位论文
源URL	[http://159.226.149.26:8080/handle/152453/10207]
专题	昆明动物研究所_真核细胞进化基因组
推荐引用方式 GB/T 7714	汪德勇. 蓝藻碳代谢途径两个关键酶FBA和FBPase的进化研究[D]. 北京. 中国科学院研究生院. 2013.

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