Biosilicification of dual-fusion enzyme immobilized on magnetic nanoparticle.
Biotechnol Bioeng
; 100(2): 223-30, 2008 Jun 01.
Article
em En
| MEDLINE
| ID: mdl-18078291
ABSTRACT
Rapid recovery, immobilization, and silica encapsulation of a dual-fusion enzyme was achieved by using iminodiacetic acid (IDA) modified magnetic nanoparticle as a carrier. D-amino acid oxidase (DAAO) of Rhodosporidium toruloides was used as a model enzyme in which a silica-precipitating peptide R5 and a metal ion complexing peptide (His)(6) were fused to its N- and C-terminal, respectively. After charging the magnetic particle with Cu(2+), the dual-fusion DAAO of 0.43 g could be directly recovered from the recombinant E. coli crude extract and immobilized on 1 g of the magnetic particle. Once in contact with hydrolyzed tetramethoxysilane (TMOS), the homogeneously dispersed immobilized dual-fusion DAAO was biosilicificated to form aggregates with size about 50 microm. The silica-encapsulated immobilized DAAO demonstrated a pyruvic acid production rate comparable with that of the naked immobilized DAAO in five repeated batch reactions when D-alanine was used as substrate. Furthermore, 85% of its activity remained after incubation at 60 degrees C for 1 h while the naked immobilized DAAO lost all its activity. This process provides the advantages that recombinant fusion enzyme can be directly recovered from crude extract, silica encapsulation protects the enzyme from leakage and denaturation, and the enzyme activity can be easily retrieved by applying a magnetic field.
Buscar no Google
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Rhodospirillum
/
Proteínas Recombinantes de Fusão
/
Dióxido de Silício
/
D-Aminoácido Oxidase
/
Enzimas Imobilizadas
/
Nanopartículas
Idioma:
En
Ano de publicação:
2008
Tipo de documento:
Article