Intensification of Organophosphorus Hydrolase Synthesis by Using Substances with Gas-Transport Function

Abstract

We have performed studies and comparative analysis of the biosynthesis characteristics of intracellular recombinant enzyme, such as hexahistidine-containing organophosphorus hydrolase (His₆-OPH) in Escherichia coli SG13009[pREP4] cells when various perfluorocarbon compounds (PFC) were introduced into the medium for cell cultivation. The PFC were found to facilitate the biosynthesis of His₆-OPH: increased levels of the total OPH-activity (up to 37%) were measured upon introduction of 1,1,1,2,2,3,3,4,4,5,5,6,6,6-tetradecafluorohexane (PFH) and 4,7,10,13,16,19,22,25,28,31-decaoxaperfluoro-5,8,11,14,17,18,21,24,27,30-decamethyl tetratriacontane (Polyether II) into culture medium. We have demonstrated the possibility of effective and multiple (at least five-fold) use of PFH for biosynthesis of intracellular recombinant protein His₆-OPH, which catalyzes the hydrolysis of organophosphorus pesticides (OP), is widely used in agriculture and can be applied as new antidote for OP-detoxification in vivo. The multiple use of PFH was achieved through recycling of this substance: sediment of Escherichia coli SG13009[pREP4] cell biomass was collected at the end of each culture growing step and disintegrated with ultrasound, and obtained residue containing almost all of the initially introduced PFC was then added to the medium at the start of the following culture growing step.