Adsorption Of Precious Metal Rh³⁺by Displaying Rhodium Metal Peptide On Cell Surface Of Pichia Pastoris

Rhodium is a precious metal with many unique properties that allow it to be widely used in industrial applications.The rapid development of modern industrial technology in China has led to significant increases in needs for rhodium,while the natural output of precious metals in China has decreased,resulting in a prominent contradiction between supply and demand for rhodium.Concomitantly,modern industrial activities continue to produce considerable amounts of rhodium-rich wastewater and waste.Consequently,rhodium recycling is an important output resource.However,conventional physical and chemical processes used in rhodium recovery exhibit many problems including high energy consumption,high cost,and serious secondary pollution.Biosorption is a novel method for treating wastewaters that contain heavy metals and has attracted considerable attention due to its high efficiency,low cost,and high level of safety.On the one hand,Pichia pastoris,as a waste biological cell in the fermentation industry,can be used to treat wastewater containing precious metals,which can greatly reduce the cost of biosorbents.On the other hand,microbial surface display technology is often used to construct new microbial adsorbents because of its ability to significantly improve the adsorption capacity and selectivity.In this study,P.pastoris and constructed recombinant P.pastoris were used as adsorbents to investigate rhodium adsorption.Several important results were observed as described below.（1）P.pastoris and Saccharomyces cerevisiae were used as adsorbents in static shake flask experiments that were conducted to investigate the effects of bacterial dosage,adsorption time,initial p H,temperature,and the co-presence of metal ions on the adsorption of Rh³⁺by two different yeast species.The optimal bacterial dosage was 0.25 g·L^-1,the optimal p H was less than or equal to 1.2,the optimal adsorption temperature was 30℃,and the optimal adsorption time required to reach maximal adsorption rates was over 2 h.Under these conditions,the equilibrium adsorption rate was 55.69%,and the equilibrium adsorption capacity was 110.1mg·g^-1.The adsorption process was most consistent with a quasi-second-order kinetic model and the Langmuir model,while thermodynamic analysis indicated that it was a monolayer chemisorption process with spontaneous endothermia.In electroplating wastewater,the maximum adsorption capacity of P.pastoris was 53.8 mg·g^-1 under optimal conditions.（2）The rhodium metal binding peptide was used as the target protein and retrieved from the Genbank database in order to synthesize its DNA sequence（Rh）.The EBY100 genome of Saccharomyces cerevisiae was also retrieved,and the pir1p sequence that is encoded as an anchor protein was obtained by PCR amplification.The recombinant plasmid p PIC9K-flag-Rh-pir1p was subsequently obtained by fusing two gene segments of the expression plasmid p PIC9K-flag-Rh-pir1p.Linear p PIC9K-flag-Rh-pir1p was then introduced into P.pastoris GS115 competent cells via electroporation to construct the recombinant P.pastoris strain GS115/p PIC9K-flag-Rh-pir1p.Recombinant P.pastoris was then induced and expressed in BMMY medium.The rhodium peptide was successfully expressed on the surface of P.pastoris cells,as determined by immunofluorescence detection.（3）The adsorption capacity of recombinant P.pastoris for Rh³⁺was studied and compared with that of waste P.pastoris.The optimal adsorption conditions of recombinant P.pastoris were consistent with that of waste P.pastoris.When the adsorption efficiency reached the maximum（75.03%）,and the equilibrium adsorption capacity reached 142.11 mg·g^-1.The adsorption process of Rh³⁺by recombinant P.pastoris was consistent with the quasi-second-order kinetic model and Temkin model.In the simulated wastewater and electroplating wastewater,the adsorption performance of the recombinant P.pastoris was 34.72%and 74.99%higher than that of the waste P.pastoris,respectively.Meanwhile,the surface display also improved the adsorption selectivity of recombinant P.pastoris.The analysis of field emission scanning electron microscopy（FESEM）and energy dispersive spectrometer（EDS）showed that a large number of nano-rhodium particles were formed on the surface of recombinant P.pastoris after adsorption.Fourier transform infrared spectrometer（FTIR）analysis showed that hydroxyl,amino,amide,carboxyl,phosphate,and sugar alcohol groups work together to adsorb Rh³⁺by recombinant P.pastoris.Desorption studies showed that 1 mol·L^-1 HNO₃ had the best desorption effect,and the desorption rate could reach 21.56%with the assistance of bacterial fragmentation.