Molecular characterization of the phaECHm, genes, required for biosynthesis of poly(3-hydroxybutyrate) in the extremely halophilic Archaeon Haloarcula marismortui
Molecular characterization of the phaECHm, genes, required for biosynthesis of poly(3-hydroxybutyrate) in the extremely halophilic Archaeon Haloarcula marismortui
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DOI：
10.1128/aem.00953-07
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发表时间：
2007-10-01
影响因子：
4.4
通讯作者：
Xiang, Hua
中科院分区：
文献类型：
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作者：
Han, Jing;Lu, Qiuhe;Xiang, Hua
虽然许多盐古菌产生可生物降解的聚羟基链烷酸酯（PHAs），但在盐古菌域中参与PRA合成的基因尚未进行实验研究。在这项研究中，我们发现，海生盐球藻能够积累聚（3-羟基丁酸酯）（PHB）细胞干重的21%，当培养在基本培养基中与过量的葡萄糖和鉴定的phaE（Hm）和phaC（Hm）基因，可能编码两个亚基的III类PHA合酶。这两个基因是相邻的，并通过一个单一的启动子位于转录起始位点上游26 bp的指导下，营养丰富和限制条件下组成型表达。有趣的是，PhaC（Hm）被揭示与PHB颗粒强烈结合，但PhaEH，m似乎不是。将phaE（Hm）或phaC（Hm）基因导入到具有高度同源性的phaEC（Hh）基因的盐囊藻中，可以提高重组菌株中的PHB合成，而两种基因的共表达总是产生最高的PHB产量。在H. hispanica导致PRA合酶活性完全丧失。与phaEC（Hm）基因的互补，但不是一个单一的，恢复的能力，聚羟基丁酸的积累，以及PHA合成酶的活性在这个phaEC缺失的盐古菌。这些结果表明，phaEC基因所需的生物合成的聚羟基丁酸，并可能编码一个活跃的PRA合酶在Haloarculd物种。
Although many haloarchaea produce biodegradable polyhydroxyalkanoates (PHAs), the genes involved in PRA synthesis in the domain of Archaea have not yet been experimentally investigated yet. In this study, we revealed that Haloarcula marismortui was able to accumulate poly (3-hydroxybutyrate) (PHB) up to 21% of cellular dry weight when cultured in a minimal medium with excessive glucose and identified the phaE(Hm) and phaC(Hm) genes, probably encoding two subunits of a class III PHA synthase. These two genes were adjacent and directed by a single promoter located 26 bp upstream of the transcriptional start site and were constitutively expressed under both nutrient-rich and -limited conditions. Interestingly, PhaC(Hm) was revealed to be strongly bound with the PHB granules, but PhaEH,m seemed not to be. Introduction of either the phaE(Hm) or phaC(Hm) gene into Haloarcula hispanica, which harbors highly homologous phaEC(Hh) genes, could enhance the PHB synthesis in the recombinant strains, while coexpression of the both genes always generated the highest PHB yield. Significantly, knockout of the phaEC(Hh) genes in H. hispanica led to a complete loss of the PRA synthase activity. Complementation with phaEC(Hm) genes, but not a single one, restored the capability of PHB accumulation as well as the PHA synthase activity in this phaEC-deleted haloarchaeon. These results indicated that the phaEC genes are required for biosynthesis of PHB and might encode an active PRA synthase in the Haloarculd species.