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Physicochemical properties of a novel α-L-arabinofuranosidase from Rhizomucor pusillus HHT-1

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Abstract:

The zygomycete fungus Rhizomucor pusillus HHT-1, cultured on L(+)arabinose as a sole carbon source, produced extracellular α-L-arabinofuranosidase. The enzyme was purified by (NH4)2SO4 fractionation, gel filtration, and ion exchange chromatography. The molecular mass of this monomeric enzyme was 88 kDa. The native enzyme had a pI of 4.2 and displayed a pH optimum and stability of 4.0 and 7.0–10.0, respectively. The temperature optimum was 65°C, and it was stable up to 70°C. The Km and Vmax for p-nitrophenyl α-L-arabinofuranoside were 0.59 mM and 387 µmol·min–1·mg–1 protein, respectively. Activity was not stimulated by metal cofactors. The N-terminal amino acid sequence did not show any similarity to other arabinofuranosidases. Higher hydrolytic activity was recorded with p-nitrophenyl α-L-arabinofuranoside, arabinotriose, and sugar beet arabinan; lower hydrolytic activity was recorded with oat–spelt xylan and arabinogalactan, indicating specificity for the low molecular mass L(+)-arabinose containing oligosaccharides with furanoside configuration.Key words: α-L-arabinofuranosidase, enzyme purification, amino acid sequence, Rhizomucor pusillus.

Le champignon zygomycète Rhizomucor pusillus HHT-1, cultivé sur du L(+)-arabinose comme seule source de carbone, a produit de la α-L-arabinofuranosidase extracellulaire. L'enzyme a été purifiée par fractionnement au (NH4)2SO4, filtration sur gel et chromatographie à échange d'ions. Le poids moléculaire de cette enzyme monomérique était de 88 kDa. L'enzyme avait un pI de 4,2, démontrait un pH optimal et une stabilité à 4,0 et 7,0–10,0, respectivement. La température optimale était de 65ºC et l'enzyme était stable jusqu'à 70°C. Le Km et le Vmax pour le p-nitrophényle α-L-arabinofuranoside étaient de 0,59 mM et 387 µmol·min–1·mg–1 de protéine, respectivement. L'activité n'a pas été stimulée par des cofacteurs métalliques. La séquence amino-terminale n'a démontré aucune similarité avec d'autres arabinofuranosidases. Une activité hydrolytique supérieure fut mesurée avec du p-nitrophényle α-L-arabinofuranoside, de l'arabinotriose et de l'arabinane de betterave à sucre, et une activité hydrolytique inférieure fut mesurée avec du xylane et de l'arabinogalactane d'épeautre d'avoine, ce qui indique une spécificité pour des oligosaccharides contenant de l'arabinose de petit poids moléculaire ayant un configuration de type furanose.Mots clés : α-L-arabinofuranosidase, purification d'enzymes, séquences d'acides aminés, Rhizomucor pusillus.[Traduit par la Rédaction]

Keywords: Rhizomucor pusillus; amino acid sequence; enzyme purification; purification d'enzymes; séquences d'acides aminés; α-L-arabinofuranosidase

Document Type: Research Article

Publication date: 2001-08-01

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  • Published since 1954, this monthly journal contains new research in the field of microbiology including applied microbiology and biotechnology; microbial structure and function; fungi and other eucaryotic protists; infection and immunity; microbial ecology; physiology, metabolism and enzymology; and virology, genetics, and molecular biology. It also publishes review articles and notes on an occasional basis, contributed by recognized scientists worldwide.
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