Evaluation Of Cu(Ⅱ) And Cr(Ⅵ) Removal Using A Metallothionein Engineered Yeast Strain

In this study, a genetically engineered strain,4126-pgk-cupl-rDNA, was achieved by integrating vectors that contained multi-copy of metallothionein gene into Saccharomyces cerevisiae4126. The tolerated and adsorption capacity of this strain was evaluated and response surface methodology was employed to optimize adsorption conditions of this strain.The metallothionein gene with native promoter,pcupl, and pgk promoter gene, as well as rDNA genes as mediated of recombinant expression were achieved by PCR amplification. Native promoter metallothionein multi-copy integration expression vector pFA6a-pcupl-rDNA and pgk promoter multi-copy integrated expression vector pFA6a-pgk-cupl-rDNA were constructed based on plasmid pFA6a using rDNA as homology arms. The vectors were transferred into S. cerevisiae4126and two engineered strains were obtained, which we proposed4126-pcupl-rDNA (4126pr) and4126-pgk-cupl-rDNA (4126pcr), respectively.The heavy metal tolerance study demonstrated that, compared with S. cerevisiae4126,4126pr displayed stronger resistance to Cu(Ⅱ), while4126pcr exhibited stronger resistance to both Cu (Ⅱ) and Cr (VI).4126pr was also proved to be more effective than S. cerevisiae4126for the adsorption of Cu (Ⅱ).4126pcr achieved an fourfold enhancement in Cu(Ⅱ) binding capacity and an improvement of40%and13%in simultaneously reduction rate of Cr(Ⅵ) and adsorption of total chromium, respectively. It was found that4126pcr exhibited a higher potential for the adsorption of heavy metals than4126pr for treating heavy metal contamination. FTIR spectra confirmed carboxyl, amino, hydroxyl group of engineered strains could be available for characteristic coordination bonding with Cu(Ⅱ), while hydroxyl, phosphate group, carboxyl, amino and amide groups were mainly involved in the adsorption process of Cr(Ⅵ) in various degrees.A quadratic model was derived for predicting the optimum conditions of removing Cr(VI) and Cu(Ⅱ) by4126pcr and the best conditions were established as fowllows:pH2,40℃adsorbent15g/L, initial concentration of Cr(VI)45mg/L and pH6,40℃, adsorbent10.44g/L, initial concentration of Cu(Ⅱ)61.75mg/L for Cr(Ⅵ) and Cu(Ⅱ), respectively. The quadratic model was verified by using the conditions mentioned above and the actual removal rates for Cr(Ⅵ) and Cu(Ⅱ) were98.35%and56.36%. which were almost consistent with the predicted value of100%and58.24%for Cr(Ⅵ) and Cu(Ⅱ), respectively. The good description of this model indicated that response surface methodology was an appropriate tool to study the absorption of Cr(Ⅵ) and Cu(Ⅱ).