Isolation of NADH from Saccharomyces cerevisiae by ether permeabilization and its purification by affinity ultrafiltration.
Indian J Exp Biol
;
1996 Oct; 34(10): 999-1004
Artigo
em Inglês
| IMSEAR
| ID: sea-58657
ABSTRACT
Cells of Saccharomyces cerevisiae were permeabilized by ether for the isolation of coenzyme NADH. A 4-fold increase in the ether fraction to aqueous fraction resulted in the recovery of 80% of total NADH present in the cell. NADH was separated and purified by affinity ultrafiltration using yeast alcohol dehydrogenase as an affinity ligand. The binding characteristics of the enzyme and coenzyme were established at different pH and ionic strengths using gel filtration. The number of moles of NADH bound per mole of alcohol dehydrogenase (r) was found to be 5.7 at pH 8 and ionic strength (I) 0.1 M. The binary complex of NADH and alcohol dehydrogenase was cleaved by lowering the pH to 6.0. The crude cell permeate on purification by ultrafiltration with 2-fold dilution, gave NADH with an absorbance ratio (A260/A340) of 2.3 and overall yield of 68%. Alcohol dehydrogenase was recovered as retentate with 93% recovery and 15% loss in activity.
Texto completo:
DisponíveL
Índice:
IMSEAR (Sudeste Asiático)
Assunto principal:
Concentração Osmolar
/
Permeabilidade
/
Saccharomyces cerevisiae
/
Álcool Desidrogenase
/
Ultrafiltração
/
Éter
/
Concentração de Íons de Hidrogênio
/
NAD
Idioma:
Inglês
Revista:
Indian J Exp Biol
Ano de publicação:
1996
Tipo de documento:
Artigo
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