A novel bacterial cell-surface display system based on NCgl1221 from Corynebacterium glutamicum
基于谷氨酸棒杆菌NCgl1221蛋白的新型细菌表面展示系统

Objective]To develop a novel Escherichia coli cell surface display system by using C-terminally truncated NCgl1221 as the anchoring protein,which greatly enriched or optimized the bacterial displayed systems.Methods]We amplified the sequence of C-terminally truncated NCgl1221 and β-amylase,and constructed the fusion expression vector.Then we transformed the recombinant plasmids PET-NA and PET-28a into Rosetta(DE3)pLysS.The fusion protein expression was induced by IPTG and identified by SDS-PAGE and Western blot analysis.The IPTG induced strains were immunostained and investigated by fluorescence microscope and flow cytometry to detect the displayed β-amylase.Finally,we analyzed the activity of β-amylase and starch hydrolization in order to determine whether the displayed β-amylase has the activity or not.Results]The fusion protein was successfully expressed in E.coli,and the active β-amylase was displayed on the cell surface by fusing it to the C terminus of the anchor.The recombinant strain displaying β-amylase can utilize soluble starch in the medium.Conclusion]A novel E.coli surface display system by using C-terminally truncated NCgl1221 as the anchor motif was successfully developed.The active enzyme with a molecular size of 56 kDa was displayed on E.coli by this system,which provided the basis for the application of the system in whole-cell biocatalyst or biosorbent.