Cloning Fuse-Expression Purification Acharacterization Of The Trehalose Synthase

Trehalose is a nonreducing disaccharide in which two glucose molecules are bonded in an a,a-1,1-glycosidic linkage, has an exceptional capacity for protecting biological membranes and proteins under stress conditions, so it shows a widely using in the future.In this paper, used nickel metal ion chromatography and Sephacryl S-300 HR gel chromatography to purify Trehalose synthase from Thermobifida fusca the recombinant enzyme with a (His)₆tag at the amino terminus and investigate properties of it. The result indication: expressed in Escherichia coli with a (His)₆ tag at the amino terminus not effect the active enzyme, Approximately 8.6mg of purified Trehalose synthase were obtained from a 1-L culture of E. coli. The recombinant enzyme showed its optimal activities at 25℃ and pH6.5, The K_m for maltose was about 15mM, whereas for trehalose it was about 311 mM.it can convert substrate 5% maltose into 60% trehalose and convert substrate 5% trehalose into 20% maltose. Active enzyme exhibited about 250kD molecular mass on PAGE...