A novel GH43 alpha-L-arabinofuranosidase from Humicola insolens: mode of action and synergy with GH51 alpha-L-arabinofuranosidases on wheat arabinoxylan.
A novel alpha-L-arabinofuranosidase (alpha-AraF) belonging to glycoside hydrolase (GH) family 43 was cloned from Humicola insolens and expressed in Aspergillus oryzae. (1)H-NMR analysis revealed that the novel GH43 enzyme selectively hydrolysed (1-->3)-alpha-L-arabinofuranosyl residues of doubly substituted xylopyranosyl residues in arabinoxylan and in arabinoxylan-derived oligosaccharides. The optimal activity of the cloned enzyme was at pH 6.7 and 53 degrees C. Two other novel alpha-L-arabinofuranosidases (alpha-AraFs), both belonging to GH family 51, were cloned from H. insolens and from the white-rot basidiomycete Meripilus giganteus. Both GH51 enzymes catalysed removal of (1-->2) and (1-->3)-alpha-L-arabinofuranosyl residues from singly substituted xylopyranosyls in arabinoxylan; the highest arabinose yields were obtained with the M. giganteus enzyme. Combinations (50:50) of the GH43 alpha-AraF from H. insolens and the GH51 alpha-AraFs from either M. giganteus or H. insolens resulted in a synergistic increase in arabinose release from water-soluble wheat arabinoxylan in extended reactions at pH 6 and 40 degrees C. This synergistic interaction between GH43 and GH51 alpha-AraFs was also evident when a GH43 alpha-AraF from a Bifidobacterium sp. was supplemented in combination with either of the GH51 enzymes. The synergistic effect is presumed to be a result of the GH51 alpha-AraFs being able to catalyse the removal of single-sitting (1-->2)-alpha-L- arabinofuranosyls that resulted after the GH43 enzyme had catalysed the removal of (1-->3)-alpha-L-arabinofuranosyl residues on doubly substituted xylopyranosyls in the wheat arabinoxylan.
Sørensen
Hanne R
HR
Novozymes A/S, Krogshøjvej 36, 2880 Bagsvaerd, Denmark.
Jørgensen
Christel T
CT
Hansen
Carsten H
CH
Jørgensen
Christian I
CI
Pedersen
Sven
S
Meyer
Anne S
AS
D000595:Amino Acid Sequence; D001089:Arabinose / Q000378:metabolism; D001203:Ascomycota / Q000201:enzymology* / Q000235:genetics; D001236:Aspergillus oryzae / Q000235:genetics; D001644:Bifidobacterium / Q000201:enzymology; D003001:Cloning, Molecular; D004795:Enzyme Stability; D015870:Gene Expression; D006026:Glycoside Hydrolases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000378:metabolism; D020072:Polyporales / Q000201:enzymology / Q000235:genetics; D016415:Sequence Alignment; D013379:Substrate Specificity; D013696:Temperature; D014908:Triticum; D014990:Xylans / Q000378:metabolism*
D009844:Oligosaccharides; D014990:Xylans; C085118:arabinoxylan; D001089:Arabinose; D006026:Glycoside Hydrolases; C019713:alpha-N-arabinofuranosidase