Covalent immobilization of acid phosphatase on amorphous AlPO4 support

Covalent attachment of acid phosphatase enzyme, AP, on the surface of amorphous AlPO4, used as inorganic support, was studied. Immobilization of the enzyme was carried out by the epsilon-amino group of lysine residues through an aromatic Schiff's-base (linker A), as well as through an 'azo' linkage to a p-OH-benzene group of tyrosine residues of the proteins (linker B). Activation of the supports in both cases was developed through the reaction of appropriate molecules with support surface -OH groups. The enzymatic activities in the 1-naphthyl phosphate hydrolysis of native, the different immobilized AP systems, and the filtrates, were obtained by a spectrophotometric method. According to the results, immobilization through linker A gave E-imm = 99% while the residual activity, E-res, at different temperatures was in the range 0.2-0.8%. On the other hand, in the immobilization by linker B, through a diazonium salt, E-imm was in the range 35-46%, but residual and specific activity values, E-res and E-spe, were between 19% and 46%. Thus, instead of linker A was more effective in the enzyme immobilization, the highest enzymatic activity after immobilization was obtained with linker B because with linker A a strong deactivation was developed. (C) 1999 Elsevier Science B.V. All rights reserved.