Simultaneous degradation of organophosphate and organochlorine pesticides by Sphingobium japonicum UT26 with surface-displayed organophosphorus hydrolase
Simultaneous degradation of organophosphate and organochlorine pesticides by Sphingobium japonicum UT26 with surface-displayed organophosphorus hydrolase
复制标题
日本鞘氨醇UT26与表面展示有机磷水解酶同时降解有机磷和有机氯农药
DOI：
10.1007/s10532-012-9587-0
复制
发表时间：
2013-04-01
期刊：
影响因子：
3.6
通讯作者：
Mulchandani, Ashok
中科院分区：
文献类型：
--
作者：
Cao, Xiangyu;Yang, Chao;Mulchandani, Ashok
通过将有机磷水解酶（OPH）基因展示在六氯环己烷（HCH）降解菌Sphingobium japonicum UT 26的细胞表面，构建了一株同时降解有机磷和有机氯农药的基因工程菌（GEM）。GEM可用于清除受污染场地混合物中可能存在的两类农药。将来自于假单胞菌的截短的冰核蛋白（INPNC）的表面锚系统用于将OPH靶向到UT 26的细胞表面上，从而降低潜在的底物摄取限制。INPNC-OPH融合的表面定位通过细胞分级分离、蛋白质印迹、蛋白酶可及性和免疫荧光显微镜进行验证。此外，通过OPH活性测定证明了表面暴露的OPH的功能性。INPNC-OPH融合蛋白（82 kDa）的表面展示既不抑制细胞生长，也不影响细胞活力。工程菌UT 26在低盐培养基中可以快速降解对硫磷和γ-HCH。研究了UT 26工程菌在无菌和非无菌土壤中对草甘膦和γ-HCH的去除效果。在这两种土壤样本中，氯磷（100 mg kg−1）和γ-六氯环己烷（10 mg kg−1）的混合物可在15天内完全降解。
A genetically engineered microorganism (GEM) capable of simultaneously degrading organophosphate and organochlorine pesticides was constructed for the first time by display of organophosphorus hydrolase (OPH) on the cell surface of a hexachlorocyclohexane (HCH)-degrading Sphingobium japonicum UT26. The GEM could potentially be used for removing the two classes of pesticides that may be present in mixtures at contaminated sites. A surface anchor system derived from the truncated ice nucleation protein (INPNC) from Pseudomonas syringae was used to target OPH onto the cell surface of UT26, reducing the potential substrate uptake limitation. The surface localization of INPNC–OPH fusion was verified by cell fractionation, western blot, proteinase accessibility, and immunofluorescence microscopy. Furthermore, the functionality of the surface-exposed OPH was demonstrated by OPH activity assays. Surface display of INPNC–OPH fusion (82 kDa) neither inhibited cell growth nor affected cell viability. The engineered UT26 could degrade parathion as well as γ-HCH rapidly in minimal salt medium. The removal of parathion and γ-HCH by engineered UT26 in sterile and non-sterile soil was also studied. In both soil samples, a mixture of parathion (100 mg kg−1) and γ-HCH (10 mg kg−1) could be degraded completely within 15 days.