title: Cloning and characteristic analysis of a novel aspartic protease gene Asp55 from Trichoderma asperellum ACCC30536.
abstract: Proteases secreted by fungi belonging to the genus Trichoderma play important roles in biocontrol. In this study, the coding sequence and promoter region of the novel aspartic protease gene Asp55 were cloned from strain Trichoderma asperellum ACCC30536. Many cis-elements involved in phytopathogenic and environmental stress responses were identified in the Asp55 promoter region and may be recognized by MYB or WRKY transcription factors. The expression pattern of Asp55 under eight culture conditions was investigated by RT-qPCR. The expression level of Asp55 was up-regulated by poplar stem powder, Alternaria alternata cell wall fragments and A. alternata fermentation liquid, while it was down-regulated by carbon and nitrogen source starvation, and by powdered poplar leaves and roots. Additionally, the expression patterns of 15 genes encoding MYB transcription factors (Myb1 to Myb15) were also analyzed by RT-qPCR. Myb2 showed the most similar expression pattern with Asp55. The cDNA of Asp55 was expressed in Escherichia coli BL21, and recombinant ASP55 (rASP55) was purified. The purified rASP55 was evaluated for enzymatic activity and showed inhibitory effect on phytopathogenic A. alternata.
authors.affiliation: School of Forestry, Northeast Forestry University, 26 Hexing Road, 150040 Harbin, China.
authors.forename: Kai
authors.initials: K
authors.lastname: Dou
authors.affiliation: School of Forestry, Northeast Forestry University, 26 Hexing Road, 150040 Harbin, China. Electronic address: wzynefu@126.com.
authors.forename: Zhiying
authors.initials: Z
authors.lastname: Wang
authors.affiliation: The College of Landscape, Northeast Forestry University, 26 Hexing Road, 150040 Harbin, China.
authors.forename: Rongshu
authors.initials: R
authors.lastname: Zhang
authors.affiliation: School of Forestry, Northeast Forestry University, 26 Hexing Road, 150040 Harbin, China.
authors.forename: Na
authors.initials: N
authors.lastname: Wang
authors.affiliation: School of Forestry, Northeast Forestry University, 26 Hexing Road, 150040 Harbin, China.
authors.forename: Haijuan
authors.initials: H
authors.lastname: Fan
authors.affiliation: School of Forestry, Northeast Forestry University, 26 Hexing Road, 150040 Harbin, China.
authors.forename: Guiping
authors.initials: G
authors.lastname: Diao
authors.affiliation: School of Forestry, Northeast Forestry University, 26 Hexing Road, 150040 Harbin, China. Electronic address: lzhnefu@126.com.
authors.forename: Zhihua
authors.initials: Z
authors.lastname: Liu
mesh_terms: D000528:Alternaria; D000595:Amino Acid Sequence; D057055:Aspartic Acid Proteases / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics; D015966:Gene Expression Regulation, Fungal; D008969:Molecular Sequence Data; D010802:Phylogeny; D060888:Real-Time Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014242:Trichoderma / Q000201:enzymology / Q000235:genetics*
chemical_list: D004271:DNA, Fungal; D005656:Fungal Proteins; D057055:Aspartic Acid Proteases
keywords: Aspartic protease ASP55; Biocontrol; Prokaryotic expression; Trichoderma asperellum