title: Heterologous overexpression and biochemical characterization of the (galactophospho)lipase from Fusarium solani in Pichia pastoris that is expressed in planta.
abstract: High-level extracellular production of Fusarium solani (galactophospho)lipase, named FSL, was achieved using a Pichia pastoris X33 expression system. The (galactophospho) lipase encoding gene was cloned into pGAPZαA with the Saccharomyces cerevisiae α-factor signal sequence by two different ways. The two constructs consist of an additional sequence of a (His)6-tag of the vector fused to the N-terminus of this enzyme (tFSL) while the other expression vector was constructed without any additional sequence (rFSL). Compared to the native enzyme (nFSL) (18.75 mg/L), a high level secretion of rFSL (310 mg/L) and tFSL (240 mg/L) was achieved providing an important improvement in enzyme production. Biochemical characterization showed that pure recombinant proteins (rFSL and tFSL) presented similar behaviour towards triglycerides, phospholipid and galactolipid. Like the nFSL, rFSL and tFSL are active at high concentration of bile salts (4mM) and calcium ions enhanced lipase activity. During plant infection, transcripts of this fungal lipase gene were detected 3, 7 and 10 days post infection.
authors.affiliation: University of SFAX, Laboratoire de Biochimie et de Génie Enzymatique des Lipases, ENIS route de Soukra, BPW 3038 Sfax, Tunisia.
authors.forename: Raida
authors.initials: R
authors.lastname: Jallouli
authors.affiliation: University of SFAX, Laboratoire de Biochimie et de Génie Enzymatique des Lipases, ENIS route de Soukra, BPW 3038 Sfax, Tunisia.
authors.forename: Madiha Bou
authors.initials: MB
authors.lastname: Ali
authors.affiliation: University of SFAX, Enzymes et Bioconversion, ENIS route de Soukra, BPW 3038 Sfax, Tunisia.
authors.forename: Mariam
authors.initials: M
authors.lastname: Charfeddine
authors.affiliation: University of SFAX, Enzymes et Bioconversion, ENIS route de Soukra, BPW 3038 Sfax, Tunisia.
authors.forename: Radhia
authors.initials: R
authors.lastname: Gargouri-Bouzid
authors.affiliation: University of SFAX, Laboratoire de Biochimie et de Génie Enzymatique des Lipases, ENIS route de Soukra, BPW 3038 Sfax, Tunisia.
authors.forename: Youssef
authors.initials: Y
authors.lastname: Gargouri
authors.affiliation: University of SFAX, Laboratoire de Biochimie et de Génie Enzymatique des Lipases, ENIS route de Soukra, BPW 3038 Sfax, Tunisia. Electronic address: sofiane_bezzine@yahoo.com.
authors.forename: Sofiane
authors.initials: S
authors.lastname: Bezzine
mesh_terms: D001647:Bile Acids and Salts / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D003001:Cloning, Molecular; D004789:Enzyme Activation / Q000187:drug effects; D004795:Enzyme Stability; D005670:Fusarium / Q000201:enzymology / Q000235:genetics*; D015870:Gene Expression; D008049:Lipase / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D010843:Pichia / Q000235:genetics*; D011994:Recombinant Proteins
chemical_list: D001647:Bile Acids and Salts; D011994:Recombinant Proteins; D008049:Lipase; D002118:Calcium
keywords: F. solani (galactophospho)lipase; Heterologous expression; P. pastoris; Phytopathology