title: Biochemical characterization, molecular cloning, and structural modeling of an interesting β-1,4-glucanase from Sclerotinia sclerotiorum.
abstract: The filamentous fungus Sclerotinia sclerotiorum produces a complete set of cellulolytic enzymes needed for efficient solubilization of native cellulose, the major component of plants. In this work, we reported the molecular characterization of an important glycosyl-hydrolase enzyme classified as endo-β-1,4-glucanase. The importance of this enzyme was revealed with the in-gel activity staining, showing a high degradation capacity of cellulose. When purified from native gel and ran in denaturing polyacrylamide gel, the polypeptide has an apparent molecular mass of about 34 kDa called Endo2. For further characterization of this protein, a mass spectrometry approach was carried out. The LC-MS/MS analysis revealed two peptides belonging to this enzyme. The genomic DNA and cDNA sequences were resolved by PCR amplification and sequencing, revealing a gene with two intron sequences. The open reading frame of 987 bp encoded a putative polypeptide of 328 amino acids having a calculated molecular mass of 33,297 Da. Yet, the molecular modeling and comparative investigation of different 3D cellulase structures showed that this endoglucanase isoform has probably two domains. A core domain having a high similarity with endoglucanases family 5 and a cellulose-binding domain having similarities with those of exo-type cellulases of family 1, linked together by a serine-threonine-rich region. These results are with great interests and show new characteristics of S. sclerotiorum glucanase.
authors.affiliation: Laboratoire d'Ingénierie des Protéines et des Molécules Bioactives (LIP-MB), National Institute of Applied Sciences and Technology, University of Carthage, 1080, Tunis Cedex, Tunisia.
authors.forename: Haifa
authors.initials: H
authors.lastname: Chahed
authors.forename: Aymen
authors.initials: A
authors.lastname: Ezzine
authors.forename: Amine Ben
authors.initials: AB
authors.lastname: Mlouka
authors.forename: Julie
authors.initials: J
authors.lastname: Hardouin
authors.forename: Thierry
authors.initials: T
authors.lastname: Jouenne
authors.forename: Mohamed Najib
authors.initials: MN
authors.lastname: Marzouki
mesh_terms: D000595:Amino Acid Sequence / Q000235:genetics; D001203:Ascomycota / Q000201:enzymology* / Q000235:genetics; D044643:Glucan 1,4-beta-Glucosidase / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D044643:Glucan 1,4-beta-Glucosidase