Subtilisin-like serine protease gene <i>TghSS42</i> from <i>Trichoderma ghanense</i> ACCC 30153 was successfully expressed in <i>Escherichia coli</i> and recombinant protease rTghSS42 exhibited antifungal ability to five phytopathogens

Abstract

The subtilisin-like serine protease gene TghSS42 was cloned from biocontrol agent Trichoderma ghanense ACCC 30153. Its coding region is 1302 bp in length, encoding 433 aa with a predicted protein molecular weight of 42.5 kDa and pI of 5.53. The accession number of cDNA sequence of TghSS42 gene is KJ740359. Furthermore, the transcription of the TghSS42 gene was all up-regulated under nine different treatments by RT-qPCR analysis, and the highest transcription level of TghSS42 approached 177.29-fold at 4 h under induction with 1% w/v Alternaria alternata cell walls, indicating that TghSS42 could be induced by the plant or phytopathogen. Furthermore, Escherichia coli recombinant strain BL21-TghSS42 was constructed. The recombinant protease rTghSS42, with an expected molecular weight of approximately 68.5 kDa containing 26.0 kDa GST tag , has been successfully expressed and purified from BL21-TghSS42. The purified protease rTghSS42 activity reached a peak of 18.7 U/mL at 4 h following 1.0 mM IPTG induction. The optimal enzyme reaction temperature was 40 and the optimal pH was 7.0. The recombinant protease rTghSS42 exerted broad-spectrum antifungal ability against Rhizoctonia solani, Fusarium oxysporum, A. alternata, Sclerotinia sclerotiorum and Cytospora chrysosperma. The inhibition rate of mycelial growth varied between 21.2% and 50.0%.

Insights into the ecological generalist lifestyle of Clonostachys fungi through analysis of their predicted secretomes

“…( Druzhinina et al, 2012 ). This family contains the serine endopeptidase subtilisin and its homologs, which have proven roles in biocontrol of fungi ( Fan et al, 2014 ; Zhang et al, 2017 ) and nematodes ( Ahman et al, 2002 ; Fekete et al, 2008 ). Numerous proteases of this class are proven to be involved in the response to mycohosts and nematodes, and this class was the most abundant one among the secreted C. rosea proteases found to be responsive to plant pathogens, together with class S1A.…”