一株降解PET单体对苯二甲酸的施氏假单胞菌的筛选和基因组分析

Screening and genome analysis of a Pseudomonas stutzeri that degrades PET monomer terephthalate

摘要:

【目的】筛选和鉴定能够降解聚对苯二甲酸乙二醇酯（PET）单体的微生物，并分析代谢途径。【方法】从青岛小涧西固体废弃物综合处置厂采集样品，以PET单体对苯二甲酸为唯一碳源筛选获得能够代谢对苯二甲酸的菌株TPA3；16S rRNA序列分析确定TPA3菌株的分类地位；采用二代和三代高通量测序技术进行基因组de novo测序和分析；通过氨基酸序列比对分析TPA3菌株代谢对苯二甲酸和乙二醇的途径和基因；验证TPA3菌株的可遗传操作性。【结果】TPA3菌株被鉴定为施氏假单胞菌（Pseudomonas stutzeri）；它能够在30℃、220 r/min培养条件下，在30 h内消耗10.60 g/L对苯二甲酸；经过驯化培养，也能够代谢乙二醇；其基因组由1条染色体和3个质粒组成，总大小约为4.55 Mb；推测该菌株具有经典的对苯二甲酸代谢途径以及与Pseudomonas putida KT2440相似的乙二醇代谢途径；采用假单胞菌遗传操作技术可对TPA3菌株进行遗传改造。【结论】TPA3菌株能够降解PET单体对苯二甲酸和乙二醇，并具有可遗传操作性，在PET废物生物处理技术中具有潜在的应用价值。

Abstract:

[Objective] To screen and identify microorganisms capable of metabolizing polyethylene terephthalate monomer terephthalate and analyze the catabolic pathways.[Methods] Collected samples from Qingdao Xiaojianxi solid waste comprehensive disposal plant and screened strains capable of metabolizing terephthalate using the medium with terephthalate as the sole carbon source; 16S rRNA phylogenetic tree analysis was used to determine the taxonomic status of TPA3 strain; de novo sequencing was implemented using the second-generation and the third-generation high-throughput sequencing technologies; the terephthalate and ethylene glycol catabolic pathways and the related genes of TPA3 were analyzed through amino acid sequence alignment; the genetic manipulability of TPA3 was verified by the reported genetic manipulative techniques.[Results] TPA3 was identified as Pseudomonas stutzeri; it can metabolize 10.60 g/L terephthalate within 30 hours at 30℃. After culture and domestication, it could also catabolize ethylene glycol. The complete genome was composed of one chromosome and three plasmids for a total genome size of 4.55 Mb. It was speculated that TPA3 strain catabolized terephthalate in a classic way, and the ethylene glycol catabolic pathway should be similar to Pseudomonas putida KT2440. The genetic manipulation technology of Pseudomonas could be used for TPA3.[Conclusion] TPA3 strain could degrade polyethylene terephthalate monomers terephthalate and ethylene glycol, and could be genetically modified, showing potential application value in polyethylene terephthalate waste biological treatment technology.