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Titolo:
Immobilization of functionally unstable catechol-2,3-dioxygenase greatly improves operational stability
Autore:
Fernandez-Lafuente, R; Guisan, JM; Ali, S; Cowan, D;
Indirizzi:
Univ Coll London, Dept Biochem & Mol Biol, London WC1E 6BT, England Univ Coll London London England WC1E 6BT Biol, London WC1E 6BT, England Univ Autonoma Madrid, CSIC, Dept Biocatalysis, Inst Catalysis, E-28049 Madrid, Spain Univ Autonoma Madrid Madrid Spain E-28049 talysis, E-28049 Madrid, Spain
Titolo Testata:
ENZYME AND MICROBIAL TECHNOLOGY
fascicolo: 8, volume: 26, anno: 2000,
pagine: 568 - 573
SICI:
0141-0229(200005)26:8<568:IOFUCG>2.0.ZU;2-B
Fonte:
ISI
Lingua:
ENG
Soggetto:
CATECHOL 2,3-DIOXYGENASE GENE; PSEUDOMONAS-PUTIDA MT-2; MULTIPOINT COVALENT IMMOBILIZATION; ESCHERICHIA-COLI; ACTIVE-SITE; STABILIZATION; CLONING; ENZYME; PURIFICATION; DEGRADATION;
Keywords:
thermophilic; thermostable; bacillus; catechol-2,3-dioxygenase; biotransformation; immobilization; stabilization;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Agriculture,Biology & Environmental Sciences
Life Sciences
Citazioni:
41
Recensione:
Indirizzi per estratti:
Indirizzo: Cowan, D Univ Coll London, Dept Biochem & Mol Biol, Gower St, London WC1E 6BT, England Univ Coll London Gower St London England WC1E 6BT E 6BT, England
Citazione:
R. Fernandez-Lafuente et al., "Immobilization of functionally unstable catechol-2,3-dioxygenase greatly improves operational stability", ENZYME MICR, 26(8), 2000, pp. 568-573

Abstract

Thermophilic catechol 2,3-dioxygenase (EC 1.13.11.2) from Bacillus stearothermophilus has been immobilized on highly activated glyoxyl agarose beads,The enzyme could be fully immobilized at 4 degrees C and pH 10.05 with a high retention of activity (around 80%). Enzyme immobilized under these conditions showed little increase in thermostability compared with the soluble enzyme, but further incubation of immobilized enzyme at 25 degrees C and pH10.05 for 3 h before borohydride reduction resulted in conjugates exhibiting a 100-fold increase in stability (c.f. the free enzyme). The stability of catechol 2,3-dioxygenase immobilized under these conditions was essentially independent of protein concentration whereas free enzyme was rapidly inactivated at low protein concentrations. An apparent stabilization factor ofover 700-fold was recorded in the comparison of free and immobilized catechol 2,3-dioxygenases at protein concentrations of 10 mu g/ml. Immobilization increased the 'optimum temperature' for activity by 20 degrees C, retained activity at substrate concentrations where the soluble enzyme was fully inactivated and enhanced the resistance to inactivation during catalysis. These results suggest that the immobilization of the enzyme under controlled conditions with the generation of multiple covalent links between the enzyme and matrix both stabilized the quaternary structure of the protein and increased the rigidity of the subunit structures. (C) 2000 Elsevier Science Inc. All rights reserved.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 07/07/20 alle ore 15:39:59