Establishment of the system for heterologous expression of catalytic subunit GLS2 of β-1,3-glucan synthase from Fusarium graminearum

This study aims to screen the heterologous expression vector of Fusarium graminearum β-1, 3-glucan synthetase catalytic subunit GLS2 (FgGLS2) and the purification detergents by Spodoptera frugiperda cell-baculovirus expression system, and will provide the basis for the study of the binding model between the protein and its inhibitor. The plasmid was constructed based on the design and modification of the baculovirus expression plasmid pFastBac by homologous recombination. The recombinant protein was heterologously expressed in the insect cell expression system and suitable detergents were screened for the extraction of the target protein. We expected to obtain the suitable vector and purification method for heterologous expression of the subunit GLS2 of β-1,3-glucan synthetase from F. graminearum. The results showed that the proteins from plasmids pFastBac-GP67-8 × His-GFP-TEV-FgGLS2, pFastBac-HA-8 × His-GFP-TEV-FgGLS2, pFastBac-GP67-6 × His-2 × Strep-TEV-FgGLS2, and pFastBac-FgRHO-TEV-8 × His could be expressed in insect cell expression system of S. frugiperda. GP67-8 × His-GFP-TEV-FgGLS2 fusion protein could be separated from cell membrane with detergent dodecyldimethylamine oxide (DDAO). HA-8 × His-GFP-TEV-FgGLS2 fusion protein can be separated from cell membrane with detergent n-dodecyl-β-maltoside(DDM), n-decyl-β-maltoside(DM) and DDAO. GP67-6 × His-2 × Strep-TEV-FgGLS2 fusion protein can be separated from cell membrane with detergent DM and DDAO. Ultimately, for the first time, we successfully express the catalytic subunit GLS2 of β-1,3-glucan synthase from F. graminearum in S. frugiperda cell expression system. In addition, the protein FgRHO could stabilize the expression of FgGLS2 and the detergent DDAO was suitable for the extraction of GLS2.