Cloning, sequence and structure of a gene encoding an antifungal glucan 1,3-beta-glucosidase from Trichoderma atroviride (T-harzianum)

A gene (gluc78) encoding an antifungal glucan 1,3-beta -glucosidase was clo ned from strain PI of the biocontrol fungus Trichoderma atroviride (formerl y T. harzianum). A putative regulatory sequence upstream from the coding re gion was cloned using single-strand extension from a primer in the known po rtion of the gene, circularized with T4 li-ase, and then reamplified with P CR to generate double-stranded DNA. The entire genomic. DNA sequence consis ted of 3440 bp, with 559 and 579 bp, respectively, in 5' and 3' untranslate d regions. The transcription unit contains a single intron, positioned in t he 5' untranslated region. The gene encodes for a protein of 770 aa, includ ing a 40 aa signal peptide. Symmetry between the first and second halves of the mature protein was found. The Gene is present as a single copy in T. a troviride and a similar gene also is present in T harzianum and T. virens. The encoded protein has similarity to a small group of sequences from filam entous fungi and no significant similarity to 1,3-beta -glucanases or gluco sidases from other organisms. Northern analysis indicates that the gene is repressed in the presence of 3% glucose and expressed in media containing 0 .1% of the sugar. Laminarin (0.1%) enhances expression after 18 h and other polymers such as scleroglucan and pustulan may enhance expression after 40 h of growth. (C) 2001 Elsevier Science B.V. All rights reserved.