PURIFICATION OF A CHITINASE FROM TRICHODERMA SP AND ITS ACTION ON SCLEROTIUM-ROLFSII AND RHIZOCTONIA-SOLANI CELL-WALLS

Trichoderma harzianum is an effective biocontrol agent of several impo rtant plant pathogenic fungi, This Trichoderma species attacks other f ungi by secreting lytic enzymes, including beta-1,3-glucanase and chit inolytic enzymes. Superior biocontrol potential may then be found in s trains having a high capacity to produce these enzymes, We have theref ore evaluated the capacity of six unidentified Trichoderma spp. isolat es to produce chitinolytic enzymes and beta-1,3-glucanases in comparis on with T. harzianum 39.1. All six isolates demonstrated substantial e nzyme activity. However, while the isolates hereafter called T-2, T-3, T-5, and T-7 produced lower amounts of enzymes, the activity of isola tes T-4 and T-6 were 2-3 fold higher than that produced by T. harzianu m 39.1. A chitinase produced by the T, isolate was purified by a singl e ion-exchange chromatography step and had a molecular mass of 46 kDa. The N-terminal amino-acid sequence showed very high homology with oth er fungal chitinases. Its true chitinase activity was demonstrated by its action on chitin and the failure to hydrolyze laminarin and p-nitr ophenyl-beta-N-acetylglucosaminide. The hydrolytic action of the purif ied chitinase on the cell wall of Sclerotium rolfsii was convincingly shown by electron microscopy studies. However, the purified enzyme had no effect on the cell wall of Rhizoctonia solani.