Main content area

Investigating and optimizing the immobilization of levansucrase for increased transfructosylation activity and thermal stability

Hill, Andrea, Karboune, Salwa, Mateo, Cesar
Process biochemistry 2017 v.61 pp. 63-72
Bacillus amyloliquefaciens, buffers, glycosides, half life, immobilized enzymes, levansucrase, pH, polyethyleneimine, prebiotics, reducing agents, response surface methodology, thermal stability
Levansucrase (LS) represents a key enzyme in glycoside synthesis of novel prebiotics and β-2,6-levan. The study of the effects of immobilization parameters of LS, produced from Bacillus amyloliquefaciens, onto glyoxyl agarose-iminodiacetic acid/Cu (glyoxyl agarose-IDA/Cu) by response surface methodology revealed the significance of their interactive effects. Retention of activity was altered by interactive effects from buffer molarity/time and buffer pH/buffer molarity. The optimized immobilization conditions were identified to be a protein loading of 9.09mg protein/g support, a buffer concentration of 608mM at pH 6.8 and an incubation time of 49h. Normally a reducing agent is applied to the immobilized enzyme in order to promote the formation of covalent bonds. This step was replaced with the addition of the ionic polymer polyethylenimine (PEI), which provided a better compromise between retained activity and thermal stability of the immobilized LS. Indeed, LS immobilized onto glyoxyl agarose-IDA/Cu/PEI had a retention of activity of 70.91% with a protein yield of 44.73% and an activity yield of 54.69%, while exhibiting a half-life 4.7 times higher than that of the free LS at 50°C.