Overproduction and properties of the mannuronate alginate lyase AlxMB |
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Authors: | Martine Malissard Frédéric Chavagnat Colette Duez Marie-Jeanne Vacheron Micheline Guinand Georges Michel Jean-Marie Ghuysen |
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Institution: | Laboratoire de Biochimie Microbienne, UniversitéClaude Bernard, 43, Boulevard du 11 Novembre 1918, F-69622 Villeurbanne Cedex, France; Centre d'Ingénierie des Protéines, Universitéde Liège, Institut de Chimie, B6, B-4000 Sart Tilman, Liège 1, Belgium |
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Abstract: | Abstract In previous studies (Malissard et al., FEMS Microbiol. Lett. (1993) 110, 101–106), the alginate lyase AlxM of the marine bacterium ATCC 433367 was produced in Escherichia coli TC4/pAL-A3 with a yield of 50 μg per litre of culture. The polypeptide chain was cleaved between two cysteine residues, C169 and C183, themselves linked by a disulphide bridge. AlxM has now been overproduced in E. coli BL21(DE3)/pAL-Sur/pLysS. Under conditions in which formation of inclusion bodies can be avoided, the enzyme is synthesized as a catalytically active, water-soluble, unnicked polypeptide with a yield of 32 mg per litre of culture. It has been purified to protein homogeneity using a one-step procedure. The nicked AlxMA and unnicked AlxMB alginate lyases have identical alginate-degrading activities at high salt concentrations. |
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Keywords: | Mannuronate alginate lyase Gene overexpression Inclusion bodies Pseudomonas aeruginosa alginate |
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