Preparation of a stable biocatalyst of bovine liver catalase using immobilization and postimmobilization techniques.
Bovine liver catalase was immobilized on different supports. The tetrameric nature of this enzyme was found to cause its rapid inactivation in diluted conditions due to subunit dissociation, a fact that may rule out its industrial use. Multi-subunit immobilization using highly activated glyoxyl agarose was not enough to involve all enzyme subunits. In fact, washing the derivative produced a strong decrease in the enzyme activity. Further cross-linking of previously immobilized enzyme with tailor-made dextran-aldehyde permitted the multimeric structure to be fully stabilized using either multisubunit preparations immobilized onto highly activated glyoxyl-agarose support or one subunit enzymes immobilized onto poorly activated glyoxyl-agarose. The highest stability of the final biocatalyst was observed using the multisubunit immobilized derivative cross-linked with dextran-aldehyde. The optimal derivative retained around 60% of the immobilized activity, did not release any enzyme subunits after boiling in the presence of SDS, and did not lose activity during washing, and its stability did not depend on the dilution. This derivative was used for 10 cycles in the destruction of 10 mM hydrogen peroxide without any decrease in the enzyme activity.
Betancor
Lorena
L
Departamento de Biocatálisis, Instituto de Catálisis, CSIC, Campus Universidad Autónoma, 28049 Madrid, Spain.
Hidalgo
Aurelio
A
Fernández-Lorente
Gloria
G
Mateo
Cesar
C
Fernández-Lafuente
Roberto
R
Guisan
José M
JM
D000818:Animals; D002374:Catalase / Q000737:chemistry*; D002384:Catalysis; D002417:Cattle; D003432:Cross-Linking Reagents / Q000737:chemistry; D004789:Enzyme Activation; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000302:isolation & purification*; D008099:Liver / Q000737:chemistry / Q000201:enzymology*; D008567:Membranes, Artificial; D011487:Protein Conformation; D011489:Protein Denaturation; D020836:Protein Structure, Quaternary; D013696:Temperature
D003432:Cross-Linking Reagents; D004800:Enzymes, Immobilized; D008567:Membranes, Artificial; D002374:Catalase