Display of both N- and C-terminal target fusion proteins on the Aspergillus oryzae cell surface using a chitin-binding module

A novel cell surface display system in Aspergillus oryzae was established by using a chitin-binding module (CBM) from Saccharomyces cerevisiae as an anchor protein. CBM was fused to the N or C terminus of green fluorescent protein (GFP) and the fusion proteins (GFP-CBM and CBM-GFP) were expressed using A. oryzae as a host. Western blotting and fluorescence microscopy analysis showed that both GFP-CBM and CBM-GFP were successfully expressed on the cell surface. In addition, cell surface display of triacylglycerol lipase from A. oryzae (tglA), while retaining its activity, was also successfully demonstrated using CBM as an anchor protein. The activity of tglA was significantly higher when tglA was fused to the C terminus than N terminus of CBM. Together, these results show that CBM used as a first anchor protein enables the fusion of both the N and/or C terminus of a target protein

Effect of Nikel on Growth and Ultrastructure of Schizosaccaromyces Pombe

In this project, we investigated effects of nickel (Ni) on the growth and ultrastructure of Schizosaccharomyces pombe wild type strain. It was found that cells were tolerant against a concentration of 2 mM Ni+2 but the generation time was extended to 5 hours from 2.5 hours for the cells growing in Ni-free YEP medium. 76% of Ni+2 was removed in 30 min by the cells grown in YEP containing 1mM Ni+2. We also analyzed the ultrastructural modifications of the cells grown in 1 mM Ni+2. There was a visible thickening of the cell wall and increase in the number of small cytoplasmic vesicles. The plasma membrane appeared irregular compared to smoother contour in control. Vacuoles contained large amounts of electron-dense materials and the size of vacuoles also increased