Kiyoshi Tagawa

Purification, Crystallization and Amino Acid Composition of α-l-Arabinofuranosidase from Aspergillus Niger

Summary 1. Crystalline α- l -arabinofuranosidase was obtained from the culture fluid of Aspergillus niger Kz. An improved method was used consisting of heat treatment, chromatography on DEAE-cellulose, DEAF-Sephadex A-50 and SE-Sephadex C-50, and gel-filtration on Sephadex G-100. 2. The molecular weight of the purified enzyme was found to be about 53 000 by the procedure of gel-filtration. The isoelectric point lies near pH 3.6. 3. The amino acid composition is shown in Table 11.

Preparation of l-arabinose-containing polysaccharides and the action of an α-l-arabinofuranosidase on these polysaccharides

Abstract A method, involving chromatography on DEAE-cellulose and gel-filtration on Sephadex G-50, is described for purification of the l -arabinan from a crude beet arabinan preparation. From the purified l -arabinan a (1 → 5)-linked l -arabinan was prepared by treatment with an α- l -arabinofuranosidase from Aspergillus niger and was characterized as a linear polymer. The l -arabinofuranose residues in these l -arabinans were hydrolyzed by action of the α- l -arabinofuranosidase, and the l -arabinose residues in wheat l -arabino- d -xylan and a small proportion of those in gum arabic were also susceptible to hydrolysis by the enzyme.

α-l-Arabinofuranosidase from Aspergillus niger

Publisher Summary α-L-Arabinofuranosidase catalyzes the hydrolysis of nonreducing terminal α-L-arabinofuranosidic linkages in L-arabinan, arabinoxylan, and other Larabinose-containing polysaccharides. In Aspergillus niger , this enzyme is produced inducibly in a medium containing L-arabinose or Larabinan. Phenyl- or p -nitrophenyl-α-L-arabinofuranoside is hydrolyzed by this enzyme action, thus, the amount of reducing group or nittrophenolate ion liberated is a measure of the enzyme activity. This chapter describes the colorimetoric assay method using p -nitrophenyl-α-L-arabinofuranoside. The chapter discusses the preparation and purification procedure of α-L-arabinofuranosidase from Aspergillus niger. The chapter concludes with a discussion on the physicochemical properties, stability, and substrate specificity of the enzyme. The enzyme is highly specific for nonreducing terminal α-L-arabinofuranosidic linkages and will not attack internal a-L-arabinofuranosyl linkages.

Isolation and some cultural conditions of Streptomyces species which produce enzymes lysing Aspergillus niger cell wall

Abstract As the result of screening microorganisms having lytic activity towards Aspergillus niger cell wall, two Streptomyces strains were isolated. One, termed J-28, was identified as Streptomyces cinereoruber and the other, termed J-13-3, was an unknown species belonging to Streptomyces . These strains inducibly produced chitinase and α-glucanase in a medium containing A. niger cell wall; specifically, strain J-28 chiefly produced chitinase and strain J-13-3 mainly α-glucanase. The best medium for chitinase production by the former was 0.5% A. niger cell wall, 0.5% yeast extract, 0.2% K 2 HPO 4 , 0.1% MgSO 4 ·7H 2 O and 0.01% FeSO 4 ·7H 2 O, while for α-glucanase production by the latter it was the same medium except for 1% instead of 0.5% A. niger cell wall, both in shaking cultures. Reaction products in chitin hydrolysis with chitinase were N , N ′-diacetyl chitobiose and chitin oligomers and those of nigeran with α-glucanase were maltose and nigeran oligomers.

Purification and properties of chitinase from Streptomyces cinereoruber

Abstract A chitinase (EC 3.2.1.14) was purified from the culture filtrate of Streptomyces cinereoruber, selected as a microorganism which produces enzymes lysing Aspergillus niger cell wall, by fractional precipitation with ammonium sulfate and column chromatographies on DEAE-cellulose, Sephadex G-100 and CM-Sephadex C-50. The final preparation was homogenous in polyacrylamide gel disc electrophoresis. The molecular weight of the enzyme was about 19,000 daltons and its isoelectric point was pH 8.6. The optimum pH and temperature for chitinase activity were 4.5 and at 50°C, respectively. The enzyme was stable in the pH range from 4.0 to 10.0. The activity was inhibited by Ag+, Hg+, Hg2+ and p-chloromercuribenzoate. Paper chromatographic analysis demonstrated that the hydrolytic products of colloidal chitin and chitotriose with the enzyme were N-acetylglucosamine and chitobiose. The lysis of A. niger cell wall with the enzyme is discussed.

Preparation of l-arabinan and 1,5-l-arabinan

Publisher Summary L-Arabinan is a polymer of L-arabinose that is found in nature wherever pectic substances occur. The molecule has a structure consisting of a chain of a-1,5-L-linked arabinofuranose units to which L-arabinofuranose units are attached mainly at position 3 in the α-configuration to form one unit side chains. 1,5-L-Arabinan is a straight chain polymer of L-arabinofuranose units derived from L-arabinan by the action of an α-L-arabinofuranosidase that preferentially splits side chains from the L-arabinan molecule. This chapter discusses the preparation of L-Arabinan and 1,5-L-Arabinan. The L-arabinan preparation is very sticky and soluble in water but insoluble in alcohols, ethers, and acetone. The 1,5-L-arabinan preparation is a white powder and insoluble in cold water but it is solubilized by heating the solution above 90°. Both the L-arabinans have a strong levorotation in aqueous solution and are easily hydrolyzed with weak acid and by a-L-arabinofuranosidases into L-arabinose, suggesting that all of the L-arabinose residues are in the furanose form and are connected by aglycosidic linkages. The chapter tabulates the properties of L-arabinan and 1,5-L-arabinan preparations obtained from beet pulp.

Studies on the Enzymes Acting on Araban: Part VIII. Purification and Properties of Arabanase Produced by Aspergillus niger

Arabanase was purified by various procedures of gel filtrations and column chromatographies from the submerged culture filtrate of Aspergillus niger grown on the medium containing beet-araban as a carbon source, and the properties of the enzyme was examined. The purified enzyme was homogeneous protein in ultracentrifugal analysis, the optimum pH for enzymatic action was 4.0. The enzyme was thermostable at pH 6.0, namely, even after heating at 98°C for 10 minutes, 20.8 per cent of the initial activity still remained. The enzyme hydrolyzed beet-araban with producing l-arabinose.

Studies on the Pectic Enzymes Part XXIII

Botrytis cinerea was cultivated in the media containing pectin, peptone, NaNO3, KH2PO4, MgSO4•7H2O and trace of FeCl3. Crude enzyme solution was prepared by salting out and dialysis, and the influence of pH value on the enzymatic action was shown. The optimum pH of endo-PG was found to be 5.4 and 3.6, and peaks of macerating action were found at pH 5.0 and 3.0 for potato disks, at pH 4.5 and 1.5 for barks of Ganpi. The macerating action at pH 4.5_??_5.0 seems to be resulted from the joint action of endo-PG and pectinesterase.