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Heterologous protein expression in pseudoalteromonas haloplanktis TAC125
No full textThe Antarctic strain Pseudoalteromonas haloplanktis TAC125 is considered one of the model organisms of cold-adapted bacteria, and during last years, it has been exploited as an alternative expression system for recombinant protein production. P. Haloplanktis TAC125 was the first Antarctic bacterium in which an efficient geneexpression technology was set up, and several generations of cold-adapted gene-expression vectors allow the production of recombinant proteins either by constitutive or inducible systems and to address the product toward any cell compartment or to the extracellular medium. Moreover, the development of synthetic media and efficient fermentation schemes, to upscale the recombinant protein production in automatic bioreactors, makes the industrial application of P. Haloplanktis TAC125 more achievable and concrete. The cellular physicochemical conditions and folding processes in P. Haloplanktis TAC125 are quite different from those observed in canonical mesophilic hosts and allowed the production of several difficult-to-express protein products, some of which are of human origin. The recently reported possibility to produce proteins within a range of temperature from 15 to -2.5 °C enhances the chances to improve the conformational quality and solubility of recombinant proteins. This chapter outlines main features and potentiality of this unconventional protein production platform
Cell engineering of Pseudoalteromonas haloplanktis TAC125: construction of a mutant strain with reduced exo-proteolytic activity
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A novel genetic system for recombinant protein secretion in the Antarctic <it>Pseudoalteromonas haloplanktis </it>TAC125
No full textAbstract Background The final aim of recombinant protein production is both to have a high specific production rate and a high product quality. It was already shown that using cold-adapted bacteria as host vectors, some "intractable" proteins can be efficiently produced at temperature as low as 4°C. Results A novel genetic system for the production and secretion of recombinant proteins in the Antarctic Gram-negative bacterium Pseudoalteromonas haloplanktis TAC125 was set up. This system aims at combining the low temperature recombinant product production with the advantages of extra-cellular protein targeting. The psychrophilic α-amylase from Pseudoalteromonas haloplanktis TAB23 was used as secretion carrier. Three chimerical proteins were produced by fusing intra-cellular proteins to C-terminus of the psychrophilic α-amylase and their secretion was analysed. Data reported in this paper demonstrate that all tested chimeras were translocated with a secretion yield always higher than 80%. Conclusion Data presented here demonstrate that the "cold" gene-expression system is efficient since the secretion yield of tested chimeras is always above 80%. These secretion performances place the α-amylase derived secretion system amongst the best heterologous secretion systems in Gram-negative bacteria reported so far. As for the quality of the secreted passenger proteins, data presented suggest that the system also allows the correct disulphide bond formation of chimera components, secreting a fully active passenger.</p
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