Biomimetic Mineralization For Enzyme Immobilization And Chemoenzymatic Cascade Catalysis

As efficient biocatalyst,natural enzymes exhibit low stability and high sensitivity to the environment,which could severely hamper their extensive application in industrial catalysis.To parallel its activity and operational stability,enzyme immobilization contributes to constructing the enzyme-carrier composites as the heterogeneous catalysts under in vitro conditions.Based on the excellent performance of enzymes in physiological environment,this work employed biomimetic mineralization materials with great biocompatibility as the carriers to confine enzymes for improving their catalytic performance and operational stability.Furthermore,the co-immobilization of cascade catalysts containing enzymes is also a significant field in enzyme immobilization.Considering that some biomimetic mineralization materials own specific catalytic activities,this work employed them as the chemical catalysts as well as carriers to build chemoenzymatic cascade catalysis systems with the aid of enzymes via one-pot method under mild conditions,which was conducive to maintaining enzyme activity and enhancing the proximity effect between enzymes and chemical catalysts,thus improving the catalysis performance of cascade catalysis systems.Hence,it provides promising opportunities to design and exploit novel chemoenzymatic cascade catalysts.Firstly,a hybrid calcium phosphate nanocrystal（CaPs）was proposed to synthesize by biomimetic mineralization method with CaPs as the inorganic component and Cu²⁺modified bovine serum albumin（BSA-Cu）as the organic component.The as-prepared hybrid composite,BSA-Cu@CaPs,was used for one-pot purification and immobilization of His-tagged organophosphorus hydrolase（OPH）.When compared to free enzyme,BSA-Cu@CaPs-OPH exhibited improved substrate（methyl parathion（MP））affinity,p H stability,and thermal stability.Moreover,BSA-Cu@CaPs-OPH could retain more than 75%and 56%of initial activity after reuse 5and 10 times,respectively.Then,Co-based zeolitic imidazolate framework（Co-ZIF）was found to be able to chemically reduce 4-nitrophenol（4-NP）to 4-aminophenol（4-AP）in the presence of NaBH₄.To enhance the tolerance of Co-ZIF against NaBH₄,we proposed a strategy of doping zinc element（Zn）in Co-ZIF to regulate the stability and catalytic activity.Then,OPH was in situ embedded under mild conditions by one-pot biomimetic mineralization method using 0.8CoZIF at a Zn²⁺/Co²⁺molar ratio of 2:8 in the initial solution as the supporting matrix.The as-obtained hybrid composite,OPH@0.8CoZIF,was employed for the chemoenzymatic cascade conversion of methyl parathion to 4-NP and then 4-AP.It could catalyze the conversion of MP at nearly 100%4-AP production within 15 min,which was 1/4 that of the physical mixed OPH and 0.8CoZIF,resulting from the proximity effect between the two catalytic activity centers in OPH@0.8CoZIF and the enrichment of MP on 0.8CoZIF.Finally,cobalt phosphate nanocrystals（CoPs）were found to exhibit catalase-like activity.Aiming at the problem that hydrogen peroxide（H₂O₂）,a catalytic by-product of partial oxidoreductases,has great damage to enzyme proteins and products,this work proposed a strategy of in-situ enzyme encapsulation of via one-pot biomimetic mineralization method using CoPs as chemical catalyst and carrier to eliminate the negative effects of H₂O₂ in time.Taking L-amino acid oxidase（LAAO）as a model enzyme,LAAO@CoPs were prepared and able to decompose H₂O₂ in time due to the high proximity of the two catalytic active centers and enhanced the proximity effect,thus avoiding the accumulation of H₂O₂ and damage on the enzyme protein and product.LAAO@CoPs could continuously and efficiently catalyze the conversion of L-tryptophan to indole-3-pyruvate with a maximum conversion more than 2 times that of LAAO system.