Heterologous expression and characterisation of the Aspergillus aspartic protease involved in the hydrolysis and decolorisation of red‐pigmented proteins
2017
Takenaka, Shinji | Umeda, Mayo | Senba, Hisanori | Koyama, Dai | Tanaka, Kōsei | Yoshida, Ken‐ichi | Doi, Mikiharu
BACKGROUND: Aspergillus repens strain MK82 produces an aspartic protease (PepA_MK82) that efficiently decolorises red‐pigmented proteins during dried bonito fermentation. However, further expansion of the industrial applications of PepA_MK82 requires the high‐level production and efficient preparation of the recombinant enzyme. RESULTS: The genomic DNA and cDNA fragments encoding the protease were cloned from strain MK82 and sequenced. Phylogenetic analysis of PepA_MK82 and comparisons with previously reported fungal aspartic proteases showed that PepA_MK 82 clusters with different groups of these enzymes. Heterologous expression of PepA_MK82 in Pichia pastoris yielded preparations of higher purity than obtained with an Escherichia coli expression system. Total protease activity in a 100‐mL culture of the P. pastoris transformant was 14 times higher than that from an equivalent culture of A. repense MK82. The recombinant PepA_MK82 was easily obtained via acetone precipitation; the final recovery was 83%. PepA_MK82 and its recombinant had similar characteristics in terms of their optimal pH, thermostability, and decolorisation activity. The recombinant was also able to decolorise flaked, dried bonito and to bleach a blood‐stained cloth. CONCLUSION: Given its ability to hydrolyse and decolorise red‐pigmented proteins, recombinant PepA_MK8 can be exploited in the food industry and as a stain‐removal agent in laundry applications. © 2016 Society of Chemical Industry
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