earticle

영어

The autodisplay technology is one of the surface display methods for proteins or peptides at the E.coli outer membrane. In the autodisplay technology, proteins or peptides are expressed as recombinant proteins in a poly protein precursor for the autotransporter secretion pathway and automatically aligned on the outer membrane surface[1]. These aligned proteins or peptides can be used as a molecular recognition layer or a affinity layer for immunoassay[2]. For the preparation of outer membrane from autodiaplyed E.coli, lysozyme was treated to hydrolyze the peptidoglycan layer and triton X-100 was used for dissolving the cell membrane. After the outer membrane isolation, marker proteins from outer membrane, periplasm, inner membrane and cytoplasm were selected and analyezed by western blot and the enzymatic assay. OmpF, β-lactamase, secA and β-galactosidase were used for western blot and KDO, β-lactamase, NADH oxidase and β -galactosidase were analyzed by enzymatic assay. Isolated outer membrane were layered on the surface of 96 well microplate by using hydrophobic interaction[3] and layered outer membrane were analyezed by enzymatic assay. The purity of isolated outer membrane and its layer was calculated and the Z-domains, have a binding affinity to Fc region of IgG, autodisplaying outer membrane layer was used for the immunoassay to improve the sensitivity.