Display of Bombyx mori alcohol dehydrogenases on the Bacillus subtilis spore surface to enhance enzymatic activity under adverse conditions

• Published in: PloS one Vol. 6; no. 6; p. e21454
• Main Authors: Wang, Nan, Chang, Cheng, Yao, Qin, Li, Guohui, Qin, Lvgao, Chen, Liang, Chen, Keping
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 29.06.2011; Public Library of Science (PLoS)
• Subjects: Adenine; Alcohol; Alcohol dehydrogenase; Alcohol Dehydrogenase - genetics; Alcohol Dehydrogenase - metabolism; Alcohol dehydrogenases; Alcohols; Aldehydes; Amylases; Animals; Antigens; Bacillus; Bacillus subtilis; Bacillus subtilis - genetics; Bacillus subtilis - metabolism; Bacillus subtilis - physiology; Biology; Bombyx - enzymology; Bombyx mori; Candida; Dehydrogenases; Drosophila melanogaster; E coli; Enzymatic activity; Enzymes; Escherichia coli; Fusion protein; Hydrogen ions; Hydrogen-Ion Concentration; Insects; Ketones; Life sciences; NADP; NADPH-diaphorase; Niacinamide; Nicotinamide; Nicotinamide adenine dinucleotide; Oxidases; Oxidation; pH effects; Phosphates; Polypeptides; Proteins; Spores; Spores, Bacterial - genetics; Spores, Bacterial - metabolism; Spores, Bacterial - physiology; Temperature; Temperature effects; Temperature range; China
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0021454

Alcohol dehydrogenases (ADHs) are oxidoreductases catalyzing the reversible oxidation of alcohols to corresponding aldehydes or ketones accompanied by nicotinamide adenine dinucleotide (NAD) or nicotinamide adenine dinucleotide phosphate (NADP) as coenzyme. ADHs attract major scientific and industrial interest for the evolutionary perspectives, afforded by their wide occurrence in nature, and for their use in industrial synthesis. However, the low activity of ADHs under extremes of pH and temperature often limits their application. To obtain ADH with high activity, in this study, we used Bombyx mori alcohol dehydrogenases (BmADH) as foreign gene and constructed a recombinant integrative plasmid pJS700-BmADH. This pJS700-BmADH was transformed into Bacillus subtilis by double cross-over and produced an amylase inactivated mutant. The fusion protein containing BmADH was expressed on the spore surface and recognized by BmADH-specific antibody. We also assayed the alcohol dehydrogenase activity of the fusion protein together with the native BmADH at different pH and temperature levels, which indicated the recombinant enzyme exhibits activity over wider ranges of temperature and pH than its native form, perhaps due to the resistance properties of B. subtilis spores against adverse conditions.