Lysyl oxidase enzyme purification and biotechnological applications

In this study, lysyl oxidase (LOX) enzyme has purified from bovine aorta by developing a new cryogel column. In the thesis, firstly methacryloyl benzotriazole (Ma-Bt) based complexes have synthesized. Then, a new chromatographic separation column has developed for LOX enzyme purification by embedding the synthesized complexes to 2-hydroxyethyl methacrylate (HEMA) based cryogel column. The synthesized complexes have characterized by Fourier Scanning Infrared (FTIR) Spectroscopy and Nuclear Magnetic Resonance (NMR) Spectroscopy. The cryogel column has characterized by FTIR, Scanning Electron Microscope (SEM) analysis and swelling tests. LOX enzyme has purified from the crude extract obtained from bovine aorta by using the synthesized cryogel as the separation column in Fast Protein Liquid Chromatography (FPLC). Then, determination of the protein content of the purified enzyme by Bradford method, activity of the purified enzyme and the optimum enzyme activity conditions have investigated. Change in the secondary and tertiary structure and the purity of the purified enzyme have investigated by Circular Dichroism (CD) spectroscopy and SDS-PAGE analysis, respectively. In this study, an effective and easily applied column material has developed for the chromatographic purification of the LOX enzyme. It has been observed that the LOX enzyme purified by the developed method is effective in cross-linking of elastin under laboratory conditions. From this point of view, it is predicted that the LOX enzyme can be used in different biotechnological applications.