Cell surface-expression of the phosphate-binding protein PstS: System development, characterization, and evaluation for phosphorus removal and recovery
Cell surface-expression of the phosphate-binding protein PstS: System development, characterization, and evaluation for phosphorus removal and recovery
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DOI：
10.1016/j.jes.2020.02.016
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发表时间：
2020-06-01
影响因子：
6.9
通讯作者：
Mayer, Brooke K.
中科院分区：
文献类型：
--
作者：
Hussein, Faten B.;Venkiteshwaran, Kaushik;Mayer, Brooke K.
无机磷(P-I)的同时过剩和稀缺是推动创新水/废水处理技术发展的一个关键问题，这些技术不仅可以促进P-I的去除以防止富营养化，而且还可以回收P-I用于农业再利用。在此，建立了细胞-表面表达的高亲和力磷酸结合蛋白(PstS)系统，并对其P-I捕获和释放潜力进行了评估。利用冰核蛋白(INP)作为锚定基序，对大肠杆菌进行基因改造，使其在其外膜上表达PstS。用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳法(SDS-PAGE)、蛋白质印迹和外膜分离分析验证蛋白质的表达和定位。通过酸碱滴定、X射线光电子能谱(XPS)、傅立叶变换红外光谱(FTIR)等手段对细胞表面进行了表征。这些测试提供了有关细菌表面的大分子结构和组成以及表面官能团的质子交换性质(即pKa值)的信息。通过磷酸盐解吸和吸附批量实验，考察了温度、pH和离子强度对磷酸盐捕集和释放的影响。与未修饰的细胞相比，PstS表面展示的细胞显示出更大的释放和捕获磷酸盐的潜力。较高的温度(40℃)、碱性pH条件(pH=10.5)和较高的离子强度(1.0mol/L KCl)促进了20%-50%的磷酸盐释放。(C)2020年中国科学院生态环境科学研究中心。爱思唯尔出版公司(Elsevier B.V.)
Simultaneous overabundance and scarcity of inorganic phosphate (P-i) is a critical issue driving the development of innovative water/wastewater treatment technologies that not only facilitate P-i removal to prevent eutrophication, but also recover P-i for agricultural reuse. Here, a cell-surface expressed high-affinity phosphate binding protein (PstS) system was developed, and its P-i capture and release potential was evaluated. E. coli was genetically modified to express PstS on its outer membrane using the ice nucleation protein (INP) as an anchoring motif. Verification of protein expression and localization were performed utilizing SDS-polyacrylamide gel electrophoresis (SDS-PAGE), western blot, and outer membrane separation analyses. Cell surface characterization was investigated through acid-base titration, X-ray photoelectron spectroscopy (XPS), and Fourier transform infrared spectroscopy (FTIR). These tests provided information on the macromolecular structure and composition of the bacteria surface as well as the proton-exchange properties of the surface functional groups (i.e., pKa values). Phosphate desorption and adsorption batch experiments were conducted to evaluate the effects of temperature, pH, and ionic strength on phosphate capture and release. The PstS surface-displayed cells demonstrated greater potential to release and capture phosphate compared to non-modified cells. Higher temperatures up to 40 degrees C, basic pH conditions (pH = 10.5), and higher ionic strength up to 1.0 mol/L KCl promoted 20%-50% higher phosphate release. (C) 2020 The Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences. Published by Elsevier B.V.