The Expression And Purification Of Recombinant Levansucrase And The Characterization And Application Of Its Product Levan

Fructan is a kind of water-soluble dietary fibre polymerized by?-D-fructopyranose.There are two primary types of fructan,inulin and levan.The digestive process of fructan and metabolic pathway are very distinct in human body,so it is of various physiological functions.In comparison with many macromolecules,the molecular weight of levan from microorganism is low.Levan from nature is white and tasteless particles with good thermal stability.The substitution solubility of levan with different polymerization degree is different.When polymerization degree decreases,the solubility will increases.According to the researchs that published,levan has low degree of polymerization,low viscosity and pesudo-plasticity.This study successfully synthesized levan from sucrose by molecular modification technology.In this paper,the enzymatic reaction process and production from sucrose of recombinant levansucrase from Erwinia amylovora ATCC49946 was studied.The gene was successfully expressed,and synthesized levan from sucrose.The research mainly studied levan from two aspects,the comparison of properties between levan and other commercial gums and the application of levan and abblications in biological composites.The optimal reaction conditions of recombinant levansucrase were pH7.0,35°C and 20%substrate concentration.Under optimized conditions,184±3 g/L levan was synthesized from500 g/L sucrose,and the conversion rate was as high as 45.96±1.25%.The molecular weight of levan was 2.017×10~7 g/mol,with uniform molecular weight distribution as the PD concentration is only 1.13±0.21.The metabolic decomposition temperature of levan from Eram-levansucrase was 216.67±4.13°C,and there was an obvious endothermic peak at125±2°C.At the concentration of 2%,levan exhibited Newtonian fluid characteristics.At 8%,levan exhibited non-newtonian pseudoplastic fluid characteristics.These results indicate that levan has great potential as microhydrogels.Therefore,the results are of great significance for the exploration of hydrogels in the food,pharmaceutical and cosmetic industries.In addition,the application of two polysaccharides-based biological composites was studied.With levan from levansucrase from Eram-levansucrase as raw material,montmorillonite and bovine serum albumin as additives,we studied the performance of Levan/MMT and Levan/BSA as biological materials.The research chosen different proportions of levan and MMT,including surface morphology observation,thermal properties and rheological behavior analysis.The results show that the thermal properties and rheological properties of Levan/MMT biocomposites were significantly improved at a ratio of 2:1.At the same time,the encapsulation rate and surface yield of 0.1%Levan/BSA nanoparticles were the highest that were 53.13±2.64%and+3.92±0.43 mV respectively.In addition,Levan/BSA nanoparticles released BSA from the system at a slower and controlled rate.These results indicated the potential applications of Levan-based biocomposites and nanoparticles.