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首页> 外文期刊>Journal of bacteriology >PhaF, a Polyhydroxyalkanoate-Granule-Associated Protein of Pseudomonas oleovorans GPo1 Involved in the Regulatory Expression System for pha Genes
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PhaF, a Polyhydroxyalkanoate-Granule-Associated Protein of Pseudomonas oleovorans GPo1 Involved in the Regulatory Expression System for pha Genes

机译：PhaF，一种油酸假单胞菌GPo1的多羟基链烷酸酯-颗粒-相关蛋白，参与pha基因的调控表达系统

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The phaC1 gene codes for the medium-chain-length polyhydroxyalkanoate (mcl PHA) synthase of Pseudomonas oleovorans GPo1, which produces mcl PHA when grown in an excess of carbon source and under nitrogen limitation. In this work, we have demonstrated, by constructing a recombinant P. oleovoransstrain carrying a phaC1::lacZreporter system, that the phaC1 gene is expressed efficiently in the presence of octanoic acid while its expression is repressed when glucose or citrate is used as the carbon source. Moreover, a P. oleovorans GPo1 mutant (strain GPG-Tc6) expressing higher levels of the reporter gene than the wild-type strain in the presence of glucose or citrate has been generated by mini-Tn5 insertional mutagenesis. Characterization of this mutant allowed us to conclude that phaF, a gene located downstream of the pha gene cluster, was knocked out in this strain. P. oleovorans GPG-Tc6 regained the ability to control phaC1 gene expression when complemented with thephaF wild-type gene. Sequencing data revealed the presence of three complete open reading frames (ORFs) in this region: ORF1 andphaI and phaF genes. The amino acid sequences of the phaI gene product and the N-terminal half of the PhaF protein showed a significant degree of similarity. Furthermore, the primary structure of the PhaF C terminus identifies this protein as a member of the histone H1-like group of proteins. Northern blot analysis showed two transcription units containing phaF, i.e., phaF and phaIF transcripts. Expression of the phaIF operon is more efficient in the presence of octanoic acid and is enhanced by the lack of the PhaF protein. In addition, it has also been demonstrated that both PhaF and PhaI proteins are bound to PHA granules produced by P. oleovorans. A model for the role of PhaF in regulating PHA synthesis is presented.
机译： phaC1 基因编码油假单胞菌 GPo1的中链长度多羟基链烷酸酯（mcl PHA）合酶，当在过量碳源下生长时产生mcl PHA。氮限制。在这项工作中，我们通过构建重组的 P证明了这一点。携带 phaC1 :: lacZ 报告系统的oleovorans 菌株，在辛酸存在时有效表达 phaC1 基因，而当葡萄糖或柠檬酸盐用作碳源时，其表达受到抑制。此外， P。 mini-Tn 5 插入诱变产生了在葡萄糖或柠檬酸存在下表达报告基因水平高于野生型菌株的oleovorans GPo1突变体（菌株GPG-Tc6）。 。对该突变体的表征使我们得出结论，该菌株中敲除了位于 pha 基因簇下游的基因 phaF 。 P。当与 phaF 野生型基因互补时，oleovorans GPG-Tc6恢复了控制 phaC1 基因表达的能力。测序数据显示该区域存在三个完整的开放阅读框（ORF）：ORF1和 phaI 和 phaF 基因。 phaI 基因产物的氨基酸序列与PhaF蛋白的N末端一半显示出显着的相似度。此外，PhaF C末端的一级结构将该蛋白鉴定为组蛋白H1样蛋白组的成员。 Northern印迹分析显示两个包含 phaF 的转录单位，即 phaF 和 phaIF 转录物。在辛酸存在下， phaIF 操纵子的表达更为有效，并且由于缺少PhaF蛋白而增强了表达。另外，还已经证明PhaF和PhaI蛋白都与 P产生的PHA颗粒结合。 oleovorans 。提出了PhaF在调节PHA合成中作用的模型。

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