High Level Expression And Immobilization Of Trehalose Synthase

It is an important way to produce trehalose by trehalose synthase,which fermentatde by Escherichia coli,catalytic maltose into trehalose.Looking for trehalose synthase which have high conversion rate,less by-product is the basis for the realization of industrialization,we can immobilize trehalose synthase and recycle to reduce the cost of industrialization.The existing laboratory-modified trehalose synthase-producing strains have been industrialized.To create greater value,on the one hand,we can immobilize the industrialized trehalose synthase.On the other hand,we can screen the better genes expressed in E.coli.Therefore,this paper mainly includes two aspects:1.The trehalose synthase was immobilized in three ways,which were cross-linked enzyme aggregates immobilization,nanoflower immobilization and resin immobilization.The cross-linked enzyme aggregates immobilization accelerates the reaction rate,in the same concentration of substrate,the reaction time is reduced by about half;The method of nanoflower immobilization is novel,and the nanoflower structure is formed.Under suitable conditions,the enzyme activity is slightly higher than free enzyme;Resin immobilization is the best method in the three methods,Low carrier price,immobilization method is simple,do not need to add cross-linking agent,can adapt to high concentrations of substrate and high temperature reaction conditions of 50?,can recycle a certain number of times.A reaction cycle is 24h,continuous use of 14 batches and the conversion rate is the 80%of the original.And the amount of by-product glucose decreased by about 3-7 g ·L-1 compared with the free enzyme.In order to overcome the shortcomings of the carrier easily broken,we carry out column reaction.A new process chart was simulated with the immobilized enzyme.After removing the by-product,and then add immobilized enzyme in the crystallized liquid to continue reacte.In this way can greatly save industrial costs.2.Five different genes were selected for expression in E.coli,and finallyscreened out a gene of Thermomonospora curvata DSM 43183.The conversion rate of the trehalose synthase in the 20 mL high concentration maltose substrate with a mass fraction of 20%was about 68.7%,which is about 14.5%higher than the laboratory.In addition,the amount of by-product glucose is reduced by about half compared to the current strain of the laboratory.From the aspects of conversion and by-products,the strain is superior to the current laboratory's.There is a great prospect of industrial applications,and it is expected to create a greater value.