Trichoderma species have been investigated as biological control agents for over 70 years owing to their ability to antagonize plant pathogenic fungi. Mycoparasitism, one of the main mechanisms involved in the antagonistic activity of Trichoderma strains, depends on the secretion of complex mixtures of hydrolytic enzymes able to degrade the host cell wall. The antifungal activity of an α‐1,3‐glucanase (EC 3.2.1.59, enzymes able to degrade α‐1,3‐glucans and also named mutanases) has been described in T. harzianum and its role in mycoparasitic processes has been suggested. In this study, we report on the purification, characterization and cloning of an exo‐α‐1,3‐glucanase, namely AGN13.2, from the antagonistic fungus T. asperellum T32. Expression at the transcription level in confrontation assays against the strawberry pathogen Botrytis cinerea strongly supports the role of AGN13.2 during the antagonistic action of T. asperellum.

The functional genomics project "TrichoEST" was developed focused on different taxonomic groups of Trichoderma with biocontrol potential. Four cDNA libraries were constructed, using similar growth conditions, from four different Trichoderma strains: Trichoderma longibrachiatum T52, Trichoderma asperellum T53, Trichoderma virens T59, and Trichoderma sp. T78. In this study, we present the analysis of the 8,160 expressed sequence tags (ESTs) generated. Each EST library was independently assembled and 1,000-1,300 unique sequences were identified in each strain. First, we queried our collection of ESTs against the NCBI nonredundant database using the BLASTX algorithm. Moreover, using the Gene Ontology hierarchy, we performed the annotation of 40.9% of the unique sequences. Later, based on the EST abundance, we examined the highly expressed genes in the four strains. A hydrophobin was found as the gene expressed at the highest level in two of the strains, but we also found that other unique sequences similar to the HEX1, QID3, and NMT1 proteins were highly represented in at least two of the Trichoderma strains.

Uricase (urate: oxygen oxidoreductase, EC 1.7.3.3) from leaves of chickpea (Cicer arietimum L.), broad bean (Vicia faba major L.), and wheat (Triticum aestivum L.) has been puri®ed to electrophoretic homogeneity by a procedure which includes xanthine-agarose anity chromatography as the main step. Puri®cation factors of 74 000±83 000 and recoveries of 80±90% were achieved. Puri®ed preparations had speci®c activities between 600 and 800 nkat á mg protein A1 (turnover numbers between 4400 and 6400 min A1 ). The three plant uricases were found by sodium dodecyl sulfate-polyacrylamide gel electrophoresis to be tetramers of similar molecular mass (120±130 kDa) and to have identical or similar-sized subunits (32±34 kDa). They also had a similar optimum pH (9±9.5) and showed a hyperbolic kinetics with K m values from 9±24 lM. All of them showed similar responses to putative activators/ inhibitors. Oxonate, xanthine and, to a lesser extent, neocuproin inhibited uricase activity, whereas allantoin, ammonium, citrulline and glutamine did not. The three leaf uricases lacked catalase activity and were not activated by cadaverine. None of the three plant enzymes cross-reacted with anti-uricase monoclonal antibodies from soybean nodules or anti-uricase polyclonal antibodies from Chlamydomonas reinhardtii or rat liver. These results are consistent with the view that uricase in plants is probably a unique enzyme which is expressed at very low level in leaves.

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Background: The filamentous fungus Trichoderma harzianum is used as biological control agent of several plant-pathogenic fungi. In order to study the genome of this fungus, a functional genomics project called "TrichoEST" was developed to give insights into genes involved in biological control activities using an approach based on the generation of expressed sequence tags (ESTs).