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Purification and characterization of endo‐xylanases from Aspergillus niger . III. An enzyme of pl 3.65
Author(s) -
Ricardo Fournier A.,
Frederick Mary M.,
Frederick James R.,
Reilly Peter J.
Publication year - 1985
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.260270422
Subject(s) - chemistry , sephadex , xylan , isoelectric point , aspergillus niger , chromatography , hydrolysis , xylanase , size exclusion chromatography , enzyme , polysaccharide , biochemistry , enzyme assay
An endo‐xylanase (1,4‐β‐ D ‐xylan xylanohydrolase, EC 3.2.1.8) from Aspergillus niger was purified to homogeneity by chromatography with Ultrogel AcA 54, SP‐Sephadex C‐25 at pH 4.5, DEAE‐Sephadex A‐25 at pH 5.4, Sephadex G‐50, and DEAE‐Sephadex A‐25 at pH 5.15. The enzyme was active on soluble xylan, on insoluble xylan only after arabinosyl‐initiated branch points were removed, and on xylooligosaccharides longer than xylotetraose. There was slight activity on carboxymethyl‐cellulose, arabinogalactan, glucomannan, and p‐nitrophenyl‐β‐ D ‐glucopyranoside. The main products of the hydrolysis of soluble and insoluble xylan were oligosaccharides of intermediate length, especially the tri‐ and pentasaccharides. The isoelectric point of the enzyme was 3.65. It had a molecular weight of 2.8 × 10 4 by SDS‐gel electrophoresis, and was high in acidic amino acids but low in those containing sulfur. Highest activity in a 20‐min assay at pH 5 was between 40 and 45°C, with an activation energy up to 40°C of 11.1 kJ/mol. The optimum pH for activity was at 5.0. The enzyme was strongly activated by Ca 2+ .