Immobilization Of D-Amino Acid Oxidase And Catalase On Muti-Walled Carbon Nanotubes

D-amino acid oxidase are widely used in D-amino acid qualitative and quantitative analysis, biosensors, L-amino acid and a-keto acid production. However, the hydrogen peroxide can be catalyzed by catalase which can promote the catalytic reaction. Immobilized enzymes have a certain influence on the catalytic reaction of the enzyme, and doublet immobilized enzymes can improve the reaction rate.In this paper, the DAAO-ELP and CAT-ELP for the research was expressed and purificated, and then, to study the immobilization of protein. It is found that the DAAO-ELP has certain stability in solution, while the dispersion is improved, the specific activity of the catalytic reaction is as much as 1.7 times than the free DAAO-ELP and the enzyme activity lasts longer. The specific activity of crosslinkde DAAO-ELP is 1.3 times of the free DAAO-ELP. In addition, by chemical crosslinking, immobilized DAAO-ELP and immobilized CAT-ELP worker toghter which has significantly increased the enzyme activity, and when there is only immobilized DAAO-ELP work the specific activity is 4 times more. And then two different proportions of immobilized enzyme obtained the different effect.The immobilized enzymes were characterized using FT-IR, CD, XPS etc. Using the UV, FT-IR, XPS spectroscopy, it is confirmed that enzyme can be immobilized. CD analysis the two structural changes of before and after the fixed enzyme, of which more than 80% alpha helix content retained, this can ensure that the immobilized enzyme has enzyme activity. In addition, through TEM, the configuration of immobilized enzyme and carrier can be observed. As to the Raman spectroscopy,it shows the change of microstructure, the enhancement of immobilized enzyme’disorder degree and the defect degree increasing.