Gene cloning of an endoglucanase from the basidiomycete Irpex lacteus and its cDNA expression in Saccharomyces cerevisiae.
A gene (cen1) coding for an endoglucanase I (En-1) was isolated from white rot fungus Irpex lacteus strain MC-2. The cen1 ORF was comprised of 399 amino acid residues and interrupted by 14 introns. The deduced amino acid sequence of the cen1 ORF revealed a multi-domain structure composed of a cellulose-binding domain, a Ser-/Thr-rich linker, and a catalytic domain from the N-terminus. It showed a significant similarity to those of other endoglucanases that belong to family 5 of glycosyl hydrolases. cen1 cDNA was inserted into a yeast expression vector, YEpFLAG-1, and introduced into Saccharomyces cerevesiae. The resulting S. cerevisiae transformant secreted a recombinant En-1 that had enzymatic properties similar to the original En-1. A strong synergistic effect for a degradation of Avicel and phosphoric acid swollen cellulose was observed when recombinant En-1 was used together with a major exo-type cellobiohydrolase I of I. lacteus MC-2.
Toda
Hiroshi
H
Department of Applied Biology, Faculty of Textile Science and Technology, Shinshu University.
Takada
Satoshi
S
Oda
Makoto
M
Amano
Yoshihiko
Y
Kanda
Takahisa
T
Okazaki
Mitsuo
M
Shimosaka
Makoto
M
D000595:Amino Acid Sequence; D001483:Base Sequence; D001487:Basidiomycota / Q000201:enzymology / Q000235:genetics*; D002480:Cellulase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002482:Cellulose / Q000378:metabolism; D043366:Cellulose 1,4-beta-Cellobiosidase / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D017422:Sequence Analysis, DNA; D020539:Sequence Analysis, Protein
D018076:DNA, Complementary; D004271:DNA, Fungal; D005656:Fungal Proteins; D011994:Recombinant Proteins; D002482:Cellulose; C056374:endoglucanase 1; D002480:Cellulase; D043366:Cellulose 1,4-beta-Cellobiosidase