NEW COMPONENTS OF THE CHITINOLYTIC SYSTEM OF TRICHODERMA-HARZIANUM

Trichoderma harzianum is a mycoparasite known as a biocontrol agent of several economically important plant pathogenic fungi. Trichoderma sp p. utilize chitinolytic enzymes, along with beta-1,3-glucanase, to deg rade the pathogen's cell walls and thus reduce disease level. We emplo yed a set of three fluorescent substrates to identify chitinolytic act ivities of proteins renatured following their separation by electropho resis. The chitinolytic system of T. harzianum was found to be more co mplex than previously reported, consisting of six distinct enzymes, tw o of which are described here for the first time. The system is compos ed of two beta-1,4 N-acetylglueosaminidases (CHIT102 and CHIT73), and four endochitinases (CHIT52, CHIT42, CHIT33 and CHIT31). The newly des cribed enzymes are CHIT73 and CHIT52. All the chitinolytic enzymes wer e induced and excreted during growth of Trichoderma on chitin as the s ole carbon source. Only CHIT102 was expressed intracellulary, at a low constitutive level, when Trichoderma was grown on glucose. Polyclonal antibodies raised against a purified 41-kDa endochitinase produced by T. harzianumm strain P1, reacted only with our CHIT42, suggesting tha t serologically all the other chitinolytic enzymes are not closely rel ated to the 41-kDa endochitinase. The complexity and diversity of the chitinolytic system of T. harzianum involves the complementary modes o f action of six enzymes, all of which are apparently required for maxi mum efficiency against a broad spectrum of chitin-containing plant pat hogenic fungi.