The bglA gene of Aspergillus kawachii encodes both extracellular and cell wall-bound beta-glucosidases.
We cloned the genomic DNA and cDNA of bglA, which encodes beta-glucosidase in Aspergillus kawachii, based on a partial amino acid sequence of purified cell wall-bound beta-glucosidase CB-1. The nucleotide sequence of the cloned bglA gene revealed a 2,933-bp open reading frame with six introns that encodes an 860-amino-acid protein. Based on the deduced amino acid sequence, we concluded that the bglA gene encodes cell wall-bound beta-glucosidase CB-1. The amino acid sequence exhibited high levels of homology with the amino acid sequences of fungal beta-glucosidases classified in subfamily B. We expressed the bglA cDNA in Saccharomyces cerevisiae and detected the recombinant beta-glucosidase in the periplasm fraction of the recombinant yeast. A. kawachii can produce two extracellular beta-glucosidases (EX-1 and EX-2) in addition to the cell wall-bound beta-glucosidase. A. kawachii in which the bglA gene was disrupted produced none of the three beta-glucosidases, as determined by enzyme assays and a Western blot analysis. Thus, we concluded that the bglA gene encodes both extracellular and cell wall-bound beta-glucosidases in A. kawachii.
Iwashita
K
K
National Research Institute of Brewing, 7-3-1, Kagamiyama, Higashihiroshima, Hiroshima 739-0046, Japan. iwasita@nrib.go.jp
Nagahara
T
T
Kimura
H
H
Takano
M
M
Shimoi
H
H
Ito
K
K
D000595:Amino Acid Sequence; D001230:Aspergillus / Q000201:enzymology* / Q000235:genetics*; D002473:Cell Wall / Q000201:enzymology; D003001:Cloning, Molecular; D005800:Genes, Fungal; D005959:Glucosidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D007438:Introns; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D001617:beta-Glucosidase / Q000737:chemistry / Q000235:genetics*
D011994:Recombinant Proteins; D005959:Glucosidases; D001617:beta-Glucosidase; C066586:6-phospho-beta-glucosidase