Functional display of active tetrameric beta-galactosidase using Bacillus subtilis spore display system.

Bum-Yeol Hwang, Jae-Gu Pan, Byung-Gee Kim, June-Hyung Kim

Department of Chemical Engineering, Bioengineering, Helen Wills Neuroscience Institute, University of California, Berkeley, CA 94720-3220, USA.

Journal of nanoscience and nanotechnology 2013 Mar

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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.

Citation

Bum-Yeol Hwang, Jae-Gu Pan, Byung-Gee Kim, June-Hyung Kim. Functional display of active tetrameric beta-galactosidase using Bacillus subtilis spore display system. Journal of nanoscience and nanotechnology. 2013 Mar;13(3):2313-9