Abstract
Monkey metallothionein α domain tandem repeats (4mMTα), which exhibit high cadmium affinity, have been displayed for the first time on the surface of a bacterium using ice nucleation protein N-domain (inaXN) protein from the Xanthomonas campestris pv (ACCC—10049) as an anchoring motif. The shuttle vector pIME, which codes for INAXN–4mMTα–EGFP fusion, was constructed and used to target 4mMTα and EGFP on the surface of Pseudomonas putida X4 (CCTCC—209319). The surface location of the INAXN–4mMTα–EGFP fusion was further verified by western blot analysis and immunofluorescence microscopy. The growth of X4 showed resistance to cadmium presence. The presence of surface-exposed 4mMTα on the engineered strains was four times higher than that of the wild-type X4. The Cd2+ accumulation by X4/pIME was not only four times greater than that of the original host bacterial cells but was also remarkably unaffected by the presence of Cu2+ and Zn2+. Moreover, the surface-engineered strains could effectively bind Cd2+ under a wide range of pH levels, from 4 to 7. P. putida X4/pIME with surface-expressed 4mMTα–EGFP had twice the cadmium binding capacity as well as 1.4 times the fluorescence as the cytoplasmic 4mMTa–EGFP. These results suggest that P. putida X4 expressing 4mMTα–EGFP with the INAXN anchor motif on the surface would be a useful tool for the remediation and biodetection of environmental cadmium contaminants.
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The research was financially supported by the Recommend International Advanced Agricultural Science & Technology Plan (“948” Program) and the National High Technology Research & Development Program of China (“863” Program, 2012AA100602).
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He, X., Chen, W. & Huang, Q. Surface display of monkey metallothionein α tandem repeats and EGFP fusion protein on Pseudomonas putida X4 for biosorption and detection of cadmium. Appl Microbiol Biotechnol 95, 1605–1613 (2012). https://doi.org/10.1007/s00253-011-3768-3
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DOI: https://doi.org/10.1007/s00253-011-3768-3