Display of oligo-α-1,6-glycosidase from <i>Exiguobacterium sibiricum</i> at the surface of <i>Escherichia coli </i>cells

Abstract

Cell-surface display based on the use of anchor motifs of outer membrane proteins allows the exposure of target peptides and proteins on the surface of microbial cells. Previously, we obtained and characterized recombinant oligo-α-1,6-glycosidase of the psychrotrophic bacterium Exiguobacterium sibiricum (EsOgl) which demonstrated high catalytic activity. It has also been shown that the autotransporter AT877 from Psychrobacter cryohalolentis and its deletion variants effectively exhibit the 10th domain of type III fibronectin (10Fn3) on the surface of Escherichia coli cells. The aim of the work was to obtain an EsOgl display system on the surface of bacterial cells based on AT877. The genes of the hybrid autotransporter EsOgl877 and its deletion mutants EsOgl877Δ239 and EsOgl877Δ310 were constructed. The enzymatic activity of EsOgl877 was investigated and it was found that the cells expressing this protein retained about 90% of the maximum activity in the range of 15-35°C. It was shown that activity of the cells containing EsOgl877Δ239 and EsOgl877Δ310 was 2.7 and 2.4 times higher, respectively, than of the cells expressing full-sized AT. Analysis of cells expressing shortened variants of EsOgl877 after treatment with proteinase K showed that the passenger domain is also localized on the cell surface. The obtained results can be used for optimization of the display systems of oligo-α-1,6-glycosidase and other heterologous proteins on the surface of E. coli cells.