Molecular cloning and sequence analysis of the cellobiohydrolase I gene from Trichoderma koningii G-39.

Abstract

The cellobiohydrolase I gene, cbh1, has been cloned from an enhanced cellulase-producing strain, Trichoderma koningii G-39. Sequence comparisons show that T. koningii cbh1 is identical to that of T. reesei with the exception of 6 bp--two causing silent substitutions in the coding region, three differing in one of the introns, and one in 5'-noncoding region. Thus, it should encode an identical CBHI to that of T. reesei despite the differences in morphological characters of the two species. Analysis of approximately 1.4 kb of the 5' flanking region shows a number of surprisingly interesting putative regulatory features. There are no unusual features within about 600 bp upstream of the translation start ATG. However, prior to the 600-bp region, there are seven CAAT sequences, a number of direct and inverted repeats, and two C/T-rich regions. Also, there are five consensus 5'-(G/C)PyGGGG-3' sequences that have been identified to be carbon catabolite repressor binding sites of Aspergillus nidulans CREA and Saccharomyces cerevisiae MIG1 repressors. The structural organization around these consensus sequence regions is similar to those of A. nidulas alcR and alcA promoters. While the production of large amounts of CBHI by T. koningii upon induction apparently correlates with the large number of CAAT boxes in the 5' upstream untranslated region of cbh1, the presence of five CREA/MIG1 repressor-binding consensus sequences in the region suggests the wide-domain carbon catabolite repression regulatory system that controls the A. nidulans ethanol regulon, and yeast GAL genes transcription might also be operative and responsible for regulation of T. koningii cbh1 transcription.