title: Cloning, molecular characterization, and mRNA expression of the thermostable family 3 β-glucosidase from the rare fungus Stachybotrys microspora.
abstract: The filamentous fungus Stachybotrys microspora possess a rich β-glucosidase system composed of five β-glucosidases. Three of them were already purified to homogeneity and characterized. In order to isolate the β-glucosidase genes from S. microspora and study their regulation, a PCR strategy using consensus primers was used as a first step. This approach enabled the isolation of three different fragments of family 3 β-glucosidase gene. A representative genomic library was constructed and probed with one amplified fragment gene belonging to family 3 of β-glucosidase. After two rounds of hybridization, seven clones were obtained and the analysis of DNA plasmids leads to the isolation of one clone (CF3) with the largest insert of 7 kb. The regulatory region shows multiple TC-rich elements characteristic of constitutive promoter, explaining the expression of this gene under glucose condition, as shown by zymogram and RT-PCR analysis. The tertiary structure of the deduced amino acid sequence of Smbgl3 was predicted and has shown three conserved domains: an (α/β)8 triose phosphate isomerase (TIM) barrel, (α/β)5 sandwich, and fibronectin type III domain involved in protein thermostability. Zymogram analysis highlighted such thermostable character of this novel β-glucosidase.
authors.affiliation: Laboratoire de Valorisation de la Biomasse et Production de Protéines chez les Eucaryotes, Centre de Biotechnologie de Sfax, University of Sfax, Route Sidi Mansour, BP 1177, 3018 Sfax, Tunisia. salma.abdeljalil@gmail.com
authors.forename: Salma
authors.initials: S
authors.lastname: Abdeljalil
authors.forename: Héla
authors.initials: H
authors.lastname: Trigui-Lahiani
authors.forename: Houcine
authors.initials: H
authors.lastname: Lazzez
authors.forename: Ali
authors.initials: A
authors.lastname: Gargouri
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015723:Gene Library; D005800:Genes, Fungal; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013191:Stachybotrys / Q000201:enzymology* / Q000235:genetics; D001617:beta-Glucosidase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D004271:DNA, Fungal; D005656:Fungal Proteins; D011994:Recombinant Proteins; D001617:beta-Glucosidase