Comparison of four phaC genes from Haloferax mediterranei and their function in different PHBV copolymer biosyntheses in Haloarcula hispanica.
Comparison of four phaC genes from Haloferax mediterranei and their function in different PHBV copolymer biosyntheses in Haloarcula hispanica.
复制标题
地中海 Haloferax 的 4 个 phaC 基因的比较及其在西班牙 Haloarcula 不同 PHBV 共聚物生物合成中的功能
DOI：
10.1186/1746-1448-6-9
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发表时间：
2010-08-20
期刊：
影响因子：
--
通讯作者：
Xiang H
中科院分区：
文献类型：
--
作者：
Han J;Li M;Hou J;Wu L;Zhou J;Xiang H
背景
嗜盐古菌Haloferax mediterranei能够从不相关的碳源中积累大量的具有高摩尔分数的3-羟基戊酸酯（3 HV）的聚（3-羟基丁酸酯-co-3-羟基戊酸酯）（PHBV）。已在该菌株中鉴定出由PhaCHme和PhaEHme两个亚基组成的聚羟基链烷酸（PHA）合酶，并显示其负责PHBV的生物合成。
结果
借助Hfx的基因组序列。2087中，鉴定了三个另外的phaC基因（命名为phaC 1、phaC 2和phaC 3），其编码推定的PhaC。与PhaCHme（54.8 kDa）一样，PhaC 1（49.7 kDa）和PhaC 3（62.5 kDa）具有III型PHA合酶的保守基序，这在PhaC 2（40.4 kDa）中没有观察到。此外，在其他三个PhaC中发现的较长的C末端在PhaC 2中也不存在。逆转录PCR（RT-PCR）结果显示，在这4个基因中，只有phaC
Hme在野生型菌株中在PHA积累条件下转录。然而，phaE的异源共表达
Hme与Haloarcula hispanica PHB-1中的每个phaC基因的比较表明，除了PhaC 2之外，所有PhaC都可以导致PHBV与各种3 HV组分的积累。采用凝胶渗透色谱（GPC）、差示扫描量热法（DSC）和热重分析（TGA）对三种共聚物进行了表征。它们的热性能随单体组成和分子量的不同而变化
w），从而可以满足各种应用要求。
结论
我们在Hfx中发现了三个隐藏的phaC基因。mediiapnei，并证明这些新鉴定的phaC基因的基因工程具有生产具有定制材料特性的PHBV的生物技术潜力。
Background
The halophilic archaeon Haloferax mediterranei is able to accumulate large amounts of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) with high molar fraction of 3-hydroxyvalerate (3HV) from unrelated carbon sources. A Polyhydroxyalkanoate (PHA) synthase composed of two subunits, PhaCHme and PhaEHme, has been identified in this strain, and shown to account for the PHBV biosynthesis.
Results
With the aid of the genome sequence of Hfx. mediterranei CGMCC 1.2087, three additional phaC genes (designated phaC1, phaC2, and phaC3) were identified, which encoded putative PhaCs. Like PhaCHme (54.8 kDa), PhaC1 (49.7 kDa) and PhaC3 (62.5 kDa) possessed the conserved motifs of type III PHA synthase, which was not observed in PhaC2 (40.4 kDa). Furthermore, the longer C terminus found in the other three PhaCs was also absent in PhaC2. Reverse transcription PCR (RT-PCR) revealed that, among the four genes, only phaC
Hme was transcribed under PHA-accumulating conditions in the wild-type strain. However, heterologous coexpression of phaE
Hme with each phaC gene in Haloarcula hispanica PHB-1 showed that all PhaCs, except PhaC2, could lead to PHBV accumulation with various 3HV fractions. The three kinds of copolymers were characterized using gel-permeation chromatography (GPC), differential scanning calorimetry (DSC), and thermogravimetric analysis (TGA). Their thermal properties changed with the variations in monomer composition as well as the different molecular weights (M
w), thus might meet various application requirements.
Conclusion
We discover three cryptic phaC genes in Hfx. mediterranei, and demonstrate that genetic engineering of these newly identified phaC genes has biotechnological potential for PHBV production with tailor-made material properties.