In recent years, a variety of extracellular proteases have been isolated from Trichoderma, most of which are serine protease and aspartic protease (table 10. 12). The molecular weight range of protease is 2 1 ~ 45 kda, with different sizes. Different Trichoderma strains can produce proteases with similar molecular weight, amino acid sequence and nucleic acid sequence. Geremia et al. (1993) isolated and purified 1 alkaline serine protease PRB 1 from Trichoderma viride, which is similar to subtilisin and similar protease from Trichoderma harzianum. In addition, 1 tryptase serine protease PRA 1 was purified from Trichoderma harzianum and Trichoderma viride, which proved that PRA 1 had nematicidal effect and could significantly reduce the hatching rate of Meloidogyne incognita eggs (Suarez et al., 2005). Trichoderma reesei can secrete acidic aspartic protease when cellulase inducer is added. It is also reported that pH-dependent aspartic protease was isolated from Trichoderma viride (Suarez et al., 2005).

Table 10. 12 Extracellular Proteases and Protease Genes from Different Trichoderma Strains

sequential

Note: n.r. was not found.

In the past few years, with people's attention to the genetic background of Trichoderma protease system, some Trichoderma protease genes have been cloned. The gene prb 1 is the first cloned Trichoderma protease gene, which encodes serine protease PRB 1. Promoter analysis showed that prb 1 contained AreA and CreA sites related to nitrogen and carbon regulation, respectively, and four heavy parasitic effector elements (MYREs)(Geremia et al., 65438+). Olmedo-Monfil et al. (2002) considered that two of these four MYREs sites were necessary for inducing prb 1 expression through parasitism. It was also found that the expression of prb 1 was inhibited by glucose, and the gene could not be expressed when nitrogen metabolism was inhibited.

Studies have shown that Trichoderma harzianum can induce protease production through mycelium or cell wall of plant pathogenic fungi. Geremia et al. (1993) isolated and identified one of the main proteases, which belongs to serine protease. Some amino acid sequences have been determined and proved to be homologous to serine protease of Bacillus subtilis protease. Poly(A)+RNA purified from Trichoderma harzianum silk was used to construct a cDNA library, and then the cDNA clone of protease was obtained by screening clones based on oligonucleotide deduced from amino acid sequence. This shows that the induced production of Trichoderma harzianum protease can be detected at RNA level. Geremia et al. (1993) isolated and identified a serine protease from Trichoderma harzianum strain IMI206040, and proved that it was related to the parasitic process of the strain, and cloned the corresponding gene prb 1. Flores et al. (1997) improved the biocontrol efficiency of Trichoderma harzianum by increasing the copy number of alkaline protease gene prb 1, and the transformant was stable, with the copy number carrying prb 1 0 being 2 ~ 10. When Trichoderma harzianum and Rhizoctonia solani interacted in vitro, prb 1 gene was expressed at a high level. Goldman( 1998) found that alkaline protease was produced during the interaction between Trichoderma harzianum and its host, and a transformed strain was constructed that overexpressed the protease.

Elad et al. (1999) found that Trichoderma harzianum T39 produced a protease, which could degrade pathogenic bacteria and digest plant cell walls, thus destroying the infectivity of botrytis cinerea. If protease inhibitor is added, the biocontrol effect of Trichoderma harzianum will be reduced. Mixing protease with Trichoderma conidia can improve the control effect. Elad et al. (1999) thought that this protease produced by Trichoderma harzianum directly inhibited the germination of pathogenic bacteria, passivated the enzymes of pathogenic bacteria and prevented the pathogenic bacteria from invading plant cells. Maria et al. (2004) cloned tvspl gene encoding extracellular serine protease from Lvmu mold and analyzed its function. It was found that the overexpression of tvspl gene could significantly improve the resistance of some Trichoderma strains to Fusarium oxysporum. Their research shows that tvspl gene is not indispensable for the normal growth of molds in Lvmu, but it plays an important role in biological control. The subtilisin prb 1 related to heavy parasitism was detected and isolated from Trichoderma echinosporus IMI206040, and the homologous gene tvsp 1 was successfully obtained from Lvmu mould Gv29.8.

Aspartic protease is one of the four main hydrolytic proteases (aspartic protease, serine protease, cysteine protease and metalloprotease), which widely exists in animals, microorganisms, viruses and plants. Their role in biological control has been confirmed (Suarez et al., 2005). Many aspartic proteases in Trichoderma have been detected and purified, and some related genes have also been cloned. The genes papA and ppB encoding Trichoderma echinosporus aspartic protease have been cloned and proved to be related to heavy parasitism (Suarez et al., 2005). Grinyer et al. (2005) successfully isolated three Aspergillus aspartic protease genes induced by Sclerotinia sclerotiorum cell wall by two-dimensional electrophoresis. Recently, Suarez et al. (2005) isolated the aspartic protease P628 1 from Trichoderma harzianum CECT24 13 and identified it as a cell wall degrading enzyme.