Sustained expression of keratinase gene under PxylA and PamyL promoters in the recombinant Bacillus megaterium MS941.
The ker gene encoding pre-pro keratinase of Bacillus licheniformis MKU3 was cloned with xylose inducible promoter (PxylA) or alpha-amylase promoter (PamyL) or both in Escherichia coli-Bacillus shuttle vector, pWH1520 generating recombinant plasmids pWHK3, pWAK3 and pWXAK3 respectively. Compared with Bacillius megaterium MS941 (pWXAK3) expressing ker gene with PxylA-PamyL promoters, B. megaterium MS941 (pWAK3) with PamyL displayed higher keratinase yield (168.6 U/ml) and specific activity (14.59 U/mg) after 36 h of growth in LB medium, however the keratinase yield decreased in the culture grown in LB medium supplemented with starch or xylose or both. A maximum yield of 186.3 U/ml with specific activity of 17.25 U/mg was obtained from xylose induced keratinase expression in B. megaterium MS941 (pWHK3) grown for 24h. The recombinant plasmids were stably maintained with sustained expression of keratinase for about 60 generations in B. megaterium MS941 rather than in B. megaterium 1,4945.
Radha
S
S
Department of Genetics, Center for Excellence in Genomic Sciences, School of Biological Sciences, Madurai Kamaraj University, Madurai 625 021, India.
Gunasekaran
P
P
D001407:Bacillus / Q000201:enzymology / Q000235:genetics; D001410:Bacillus megaterium / Q000201:enzymology* / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics; D003001:Cloning, Molecular; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D006090:Gram-Negative Bacteria / Q000201:enzymology / Q000235:genetics; D010447:Peptide Hydrolases / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000378:metabolism; D000516:alpha-Amylases / Q000235:genetics*
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D000516:alpha-Amylases; D010447:Peptide Hydrolases; C035855:keratinase