Molecular cloning and overexpression of fleA gene encoding a fucose-specific lectin of Aspergillus oryzae.
A protein from the cell lysate of Aspergillus oryzae was purified by column chromatography immobilized with a ferrichrysin (Fcy), which is one of the siderophores of A. oryzae. It is produced only in an iron-deficient culture and its molecular weight is estimated as 35,000 by SDS-PAGE. Two internal amino acid sequences of the protein obtained by lysylendopeptidase digestion were analyzed. Molecular cloning shows that it encodes 310 putative amino acid residues separated by 4 introns and is designated as fleA. It shows approximately 26% similarity with the gene encoding a fucose-specific lectin of Aleuria aurantia (AAL). The gene was overexpressed under control of the melO promoter in a submerged culture of A. oryzae. The fleA gene product showed hemagglutination activity against rabbit erythrocytes. A hemagglutination inhibition assay of monosaccharides showed that this lectin specifically binds to L-fucose and weakly reacts with mannose and N-acetyl-neuraminic acid.
Ishida
Hiroki
H
Research Institute, Gekkeikan Sake Co. Ltd, Kyoto, Japan. h-ishida@gekkeikan.co.jp
Moritani
Toshiyasu
T
Hata
Yoji
Y
Kawato
Akitsugu
A
Suginami
Koji
K
Abe
Yasuhisa
Y
Imayasu
Satoshi
S
D000595:Amino Acid Sequence; D001236:Aspergillus oryzae / Q000235:genetics*; D001483:Base Sequence; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004271:DNA, Fungal; D004591:Electrophoresis, Polyacrylamide Gel; D005643:Fucose / Q000378:metabolism*; D037102:Lectins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction
D004271:DNA, Fungal; D037102:Lectins; D005643:Fucose