title: Functional display of active tetrameric beta-galactosidase using Bacillus subtilis spore display system.
abstract: For the functional bacterial surface display of active enzyme of multimeric form, which is generally impossible due to molecular assembly of the monomer subunit subsequent to the secretion of displayed target protein outside the cell, a new surface display system based on B. subtilis spore was developed. Using cotE and cotG of B. subtilis as anchoring motives, beta-galactosidase, which is active in tetrameric form, was functionally displayed on the surface of B. subtilis spore. The surface localization of beta-galactosidase was verified by Miller assay of purified spore, protease accessibility test of purified spore, and flow cytometric analysis of spore expressing beta-galactosidase. While B. subtilis spore wall integrity, examined by lysozyme and heat treatments, was affected by the incorporation of CotE-LacZ fusion protein, it was not affected by the incorporation of CotG-lacZ fusion. Heat stability of displayed protein was similar with that of free enzyme.
authors.affiliation: Department of Chemical Engineering, Bioengineering, Helen Wills Neuroscience Institute, University of California, Berkeley, CA 94720-3220, USA.
authors.forename: Bum-Yeol
authors.initials: BY
authors.lastname: Hwang
authors.forename: Jae-Gu
authors.initials: JG
authors.lastname: Pan
authors.forename: Byung-Gee
authors.initials: BG
authors.lastname: Kim
authors.forename: June-Hyung
authors.initials: JH
authors.lastname: Kim
mesh_terms: D001412:Bacillus subtilis / Q000254:growth & development*; D001483:Base Sequence; D001704:Biopolymers / Q000737:chemistry*; D017931:DNA Primers; D005434:Flow Cytometry; D006358:Hot Temperature; D009113:Muramidase / Q000737:chemistry; D013171:Spores, Bacterial; D001616:beta-Galactosidase / Q000737:chemistry*
chemical_list: D001704:Biopolymers; D017931:DNA Primers; D009113:Muramidase; D001616:beta-Galactosidase