Purification of the Early Sporulation Gene spo0F Product of Bacillus subtilis |
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Authors: | Masanao Oda Masami Isaka Yoshikatsu Murooka Ryosaku Nomi Hirofumi Yoshikawa Fujio Kawamura |
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Institution: | 1. Department of Applied Biochemistry, Hiroshima University, Fukuyama 720, Japan;2. Department of Fermentation Technology, Hiroshima University, Higashihiroshima 724, Japan;3. Institute of Applied Microbiology, University of Tokyo, Tokyo 113, Japan |
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Abstract: | The RsaI fragment (750 base pairs) containing the entire early sporulation gene spo0F of Bacillus subtilis was inserted in the downstream region of the PR promoter of the expression vector, pEBR-151. The recombinant plasmid thus obtained was introduced into Escherichia coli HB101 and the synthesis of the spo0F gene product was induced by a temperature shift up. After induction for 7 hr, a protein of molecular weight 14,000 (14 K protein) was overproduced to about 9% of the total cellular protein. The 14 K protein was purified to 94% purity by four steps of column chromatography. Deletion analysis and the sequence determination of the NH2-terminal amino acid residues of the purified 14 K protein confirmed that the 14 K protein is the spo0F gene product. |
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Keywords: | chitosanase Paenibacillus fukuinensis family 8 glycosyl hydrolase yeast cell-surface engineering |
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