Functional cloning of an endo-arabinanase from Aspergillus aculeatus and its heterologous expression in A. or oryzae and tobacco.
Functional cloning in yeast has been used to isolate full-length cDNAs encoding an endo-alpha-1,5-L-arabinanase from the filamentous fungus Aspergillus aculeatus. Screening of a cDNA library constructed in a yeast expression vector for transformants that hydrolysed AZCL-arabinan identified 44 Saccharomyces cerevisiae clones all harbouring the same arabinanase-encoding cDNA. The cloned cDNA was expressed in A. oryzae and the recombinant enzyme was purified and characterized. The mode of action of the enzyme was studied by analysis of the digestion pattern towards debranched arabinan. The digestion profile obtained strongly suggests that the enzyme is an endo-arabinanase. In addition, the feasibility using Nicotiana tabacum as an alternative host for arabinanase expression was investigated.
Skjøt
M
M
Biotechnology Group, Danish Institute of Agricultural Sciences, Copenhagen. m.skjot@dias.kvl.dk
Kauppinen
S
S
Kofod
L V
LV
Fuglsang
C
C
Pauly
M
M
Dalbøge
H
H
Andersen
L N
LN
D000595:Amino Acid Sequence; D001230:Aspergillus / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D005800:Genes, Fungal; D006026:Glycoside Hydrolases / Q000235:genetics*; D008969:Molecular Sequence Data; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D016415:Sequence Alignment; D014026:Nicotiana / Q000235:genetics
D006026:Glycoside Hydrolases; C019713:alpha-N-arabinofuranosidase