Lignin degradation, ligninolytic enzymes activities and exopolysaccharide production by Grifola frondosa strains cultivated on oak sawdust

Fourteen strains of Grifola frondosa (Dicks.) S. F. Gray, originating from different regions (Asia, Europe and North America) were tested for lignin degradation, ligninolytic enzyme activities, protein accumulation and exopolysaccharide production during 55 days of cultivation on oak sawdust. Lignin degradation varied from 2.6 to7.1 percent of dry weight of the oak sawdust substrate among tested strains. The loss of dry matter in all screened fungi varied between 11.7 and 33.0 percent, and the amount of crude protein in the dry substrate varied between 0.94 to 2.55 percent. The strain, MBFBL 596, had the highest laccase activity (703.3 U/l), and the maximum peroxidase activity of 22.6 U/l was shown by the strain MBFBL 684. Several tested strains (MBFBL 21, 638 and 662) appeared to be good producers of exopolysaccharides (3.5, 3.5 and 3.2 mg/ml respectively).

Molecular cloning, expression and characterization of a serine proteinase from Japanese edible mushroom, Grifola frondosa: solving the structure - function anomaly of a reported aminopeptidase

The N-terminal amino acid sequence of an aminopeptidase from Japanese edible mushroom, Grifola frondosa, was reported to have high similarity with that of a serine proteinase from basidiomycete, Agaricus bisporous (Nishiwaki and Hayashi, 2001). The full-length cDNA and the corresponding genomic DNA of the enzyme were cloned, based on the reported N-terminal amino acid sequence. The predicted open reading frame (ORF) of the cloned cDNA, encoding a product of 379 amino acids, was expressed in E. coli using pET expression vector. The expressed pro-enzyme (40 kDa) underwent autolysis to produce the mature protein (30 kDa) and a pro-peptide (10 kDa). The mature protein and the pro-peptide remained tightly bound to each other and could not be separated by Ni-NTA metal affinity chromatography or Q-Sepharose ion-exchange chromatography. The enzyme was inactive in the bound form. Upon treatment with subtilisin, the bound pro-peptide was further hydrolyzed and a high serine proteinase activity was recovered. No aminopeptidase activity was detected at any stage of the protein processing. These results clearly indicated that the N-terminal amino acid sequence and the function of the reported aminopeptidase were not derived from the same protein entity and hence caused the structure-function anomaly.