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Titolo:
RENATURATION, PURIFICATION, AND CHARACTERIZATION OF RECOMBINANT D-2-HYDROXYISOCAPROATE DEHYDROGENASE FROM ESCHERICHIA-COLI
Autore:
AGRAZ A; PAULSEN J; BORNER B; HUSTEDT H;
Indirizzi:
GESELL BIOTECHNOL FORSCH MBH,MASCHERODER WEG 1 D-38124 BRAUNSCHWEIG GERMANY GESELL BIOTECHNOL FORSCH MBH D-38124 BRAUNSCHWEIG GERMANY CTR INGN GENET & BIOTECNOL HAVANA CUBA
Titolo Testata:
Enzyme and microbial technology
fascicolo: 6, volume: 17, anno: 1995,
pagine: 558 - 563
SICI:
0141-0229(1995)17:6<558:RPACOR>2.0.ZU;2-M
Fonte:
ISI
Lingua:
ENG
Soggetto:
LACTOBACILLUS-CASEI; STEREOSPECIFIC REDUCTION; 2-KETOCARBOXYLIC ACIDS; PROTEINS; ENZYME;
Keywords:
RECOMBINANT PROTEIN; D-2-HYDROXYISOCAPROATE DEHYDROGENASE; INCLUSION BODIES; RENATURATION;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
20
Recensione:
Indirizzi per estratti:
Citazione:
A. Agraz et al., "RENATURATION, PURIFICATION, AND CHARACTERIZATION OF RECOMBINANT D-2-HYDROXYISOCAPROATE DEHYDROGENASE FROM ESCHERICHIA-COLI", Enzyme and microbial technology, 17(6), 1995, pp. 558-563

Abstract

Conditions for renaturation and purification of recombinant D-2-hydroxyisocaproate dehydrogenase (r-DHicDH) produced in E. coli in the formof inclusion bodies were investigated. Urea (8 M) was found to extract the enzyme from the pellet fraction efficiently. Renaturation by dialysis against 20 mM sodium phosphate buffer, pH 7.0, a protein concentration of 2 mg ml(-1), and a temperature of 4-8 degrees C was found tobe optimal and even at a protein concentration of 10 mg ml(-1) 85% ofenzyme activity could be recovered. Protein was further purified by ion-exchange chromatography on a Mono Q column. An enzyme preparation of greater than 95% purity was obtained with an overall yield of about 50%. r-DHicDH shows identical properties within experimental error with respect to molecular weight, temperature dependence of enzyme activity, pH optimum of the enzyme reaction, and K-M values for 2-ketoisocaproate and NADH as the native enzyme from Lactobacillus casei.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 21/09/20 alle ore 04:43:06