Heterologous overexpression and biochemical characterization of the (galactophospho)lipase from Fusarium solani in Pichia pastoris that is expressed in planta

• A high level expression of a novel (phospho)lipase from F. solani was achieved.
• The histidine extension of the recombinant enzyme does not affect its properties.
• Possible involvement of this enzyme in the phytopathological process.

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 (4 mM) and calcium ions enhanced lipase activity.During plant infection, transcripts of this fungal lipase gene were detected 3, 7 and 10 days post infection.