Cloning and expression of a novel Trichoderma viride laminarinase AI gene (lamAI).
The gene lamAI, which encodes a novel laminarinase AI of Trichoderma viride U-1, was cloned using RT-PCR in conjunction with the rapid amplification of cDNA ends (RACE) technique. The open reading frame consisted of 2,277 bp encoding a protein of 759 amino acid residues, including a 32-residue signal prepropeptide. The protein showed 91% sequence similarity to the putative Trichoderma virens beta-1,3-glucanase BGN1, but no significant similarity to fungal beta-1,6-glucanases or beta-1,3-glucanases from other organisms. On 40 h incubation with a solo carbon source, northern analysis revealed that the gene was induced by 0.5% laminaran from Eisenia bicyclis but was not by the same concentration of glucose. The lamAI cDNA was functionally expressed in the methylotrophic yeast Pichia pastoris, resulting in a recombinant enzyme with as high activity against laminaran as native LAMAI. Based on these data, the probable existence of endo-beta-1,3:1,6-glucan hydrolases as a subclass of endo-beta-1,3-glucanases in some mycoparasitic fungi is suggested.
Nobe
Rika
R
Department of Biochemistry and Applied Biosciences, Faculty of Agriculture, University of Miyazaki, Miyazaki 889-2192, Japan.
Sakakibara
Yoichi
Y
Ogawa
Kihachiro
K
Suiko
Masahito
M
D000595:Amino Acid Sequence; D001483:Base Sequence; D044602:Cellulases / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal / Q000235:genetics*; D005936:Glucans; D005947:Glucose / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics / Q000378:metabolism; D011134:Polysaccharides / Q000378:metabolism*; D014242:Trichoderma / Q000235:genetics* / Q000378:metabolism
D018076:DNA, Complementary; D005656:Fungal Proteins; D005936:Glucans; D011134:Polysaccharides; C008247:laminaran; D044602:Cellulases; D006026:Glycoside Hydrolases; D005947:Glucose