Cell engineering of Pseudoalteromonas haloplanktis TAC125: construction of a mutant strain with reduced exo-proteolytic activity

Background We have already shown that using cold-adapted bacteria as host vectors, some "intractable" proteins can be efficiently produced at temperature as low as 4°C [1,2]. Furthermore, we set up a "cold" gene-expression system implemented for the secretion of recombinant proteins in the Antarctic Gram-negative bacterium Pseudoalteromonas haloplanktis TAC125 (PhTAC125). Such a system could effectively conjugate the positive effect of low temperature on the recombinant product solubility with the obvious advantages linked to extra-cellular protein targeting. This novel system makes use of the psychrophilic α-amylase from PhTAB23 [3] as secretion carrier. Several chimerical proteins were produced and used to test the versatility and efficiency of the novel secretion system. All the chimerical proteins were efficiently produced and secreted (Cusano AM, Ph. D thesis 2005 Università di Napoli "Federico II"). However, bacteria belonging to Pseudoalteromonas genus are reported to secrete a wide range of exo-proteins, especially proteases. This feature could hamper both applicability and efficiency of the cold-adapted secretion system, due to the possible recombinant product degradation.