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 cloned from strain P1 of the biocontrol fungus Trichoderma atroviride (formerly T. harzianum). A putative regulatory sequence upstream from the coding region was cloned using single-strand extension from a primer in the known portion of the gene, circularized with T4 ligase, and then reamplified with PCR to generate double-stranded DNA. The entire genomic DNA sequence consisted of 3440 bp, with 559 and 579 bp, respectively, in 5' and 3' untranslated regions. The transcription unit contains a single intron, positioned in the 5' untranslated region. The gene encodes for a protein of 770 aa, including 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. atroviride 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 filamentous fungi and no significant similarity to 1,3-beta-glucanases or glucosidases 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.
Donzelli
B G
BG
Department of Horticultural Sciences, Cornell University, Geneva, NY 14456, USA. bdd1@cornell.edu
Lorito
M
M
Scala
F
F
Harman
G E
GE
D024506:5' Flanking Region / Q000235:genetics; D000595:Amino Acid Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D005091:Exons; D005658:Fungi / Q000201:enzymology; D015971:Gene Expression Regulation, Enzymologic; D015966:Gene Expression Regulation, Fungal; D005796:Genes / Q000235:genetics*; D043326:Glucan 1,3-beta-Glucosidase; D007438:Introns; D008969:Molecular Sequence Data; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D024363:Transcription Initiation Site; D014242:Trichoderma / Q000201:enzymology / Q000235:genetics*; D001617:beta-Glucosidase / Q000235:genetics*
D004271:DNA, Fungal; D012331:RNA, Fungal; D001617:beta-Glucosidase; D043326:Glucan 1,3-beta-Glucosidase