AIMS Bioengineering, 2017, 4(3): 386-401. doi: 10.3934/bioeng.2017.3.386.

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Identification, recombinant production and partial biochemical characterization of an extracellular cold-active serine-metalloprotease from an Antarctic Pseudomonas isolate

1 Biochemistry and Molecular Biology, Faculty of Sciences, Universidad de la República, Iguá 4225, Montevideo, Uruguay
2 Molecular Microbiology, Institute Clemente Estable, Av. Italia 3318, Montevideo, Uruguay
3 Unité de Biochimie Structurale, Institut Pasteur, URA-CNRS 2185, 75724 Paris cedex 15, France

Cold-adapted enzymes are generally derived from psychrophilic microorganisms and have features that make them very attractive for industrial and biotechnological purposes. In this work, we identified a 50 kDa extracellular protease (MP10) from the Antarctic isolate Pseudomonas sp. AU10. The enzyme was produced by recombinant DNA technology, purified using immobilized metal affinity chromatography and partially characterized. MP10 is an alkaline thermosensitive serine-metallo protease with optimal activity at pH 8.0 and 40 ℃, in the presence of 1.5 mM Ca2+. MP10 showed 100% residual activity and stability (up to 60 min) when incubated with 7% of non-ionic surfactants (Triton X-100, Tween-80 and Tween-20) and 1.5% of the oxidizing agent hydrogen peroxide. The 3D MP10 structure was predicted and compared with the crystal structure of mesophilic homologous protease produced by Pseudomonas aeruginosa PA01 (reference strain) and other proteases, showing similarity in surface area and volume of proteins, but a significantly higher surface pocket area and volume of MP10. The observed differences presumably may explain the enhanced activity of MP10 for substrate binding at low temperatures. These results give insight to the potential use of MP10 in developing new biotechnologically processes active at low to moderate temperatures, probably with focus in the detergent industry.
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Keywords protease; Pseudomonas; cold-adaptation; surfactant; Antarctica

Citation: Natalia Fullana, Victoria Braña, Juan José Marizcurrena, Danilo Morales, Jean-Michel Betton, Mónica Marín, Susana Castro-Sowinski. Identification, recombinant production and partial biochemical characterization of an extracellular cold-active serine-metalloprotease from an Antarctic Pseudomonas isolate. AIMS Bioengineering, 2017, 4(3): 386-401. doi: 10.3934/bioeng.2017.3.386


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