Second University of Naples
Title: Fermentation strategies for recombinant thermophilic phosphotriesterase enzyme production
Odile Francesca Restaino has got a Master Degree in Organic Chemistry at the “Federico II” University in Naples and completed her Ph.D. in 2008 at the Second University of Naples in biotechnology field. She got a Fulbright Research Fellowship in 2012, as visiting scientist to work in the “Bioengineered heparin project” in prof. Linhardt lab at Rensselaer Polytechnic Institute, NY, USA. Her background includes analytical chemistry, fermentation process development and downstream purification strategies for structural characterization and biotechnological production of bio-molecules for industrial applications and of natural-like drugs as chondroitin sulfate and heparin.
The organophosphate (OP) are widely used as insecticides and some of them could be used as nerve agents. Thus the enzymes able to degrade this class of compounds resulted of great interest from a scientific and industrial point of view because they could eventually be used as de-activated agent. A thermostable phosphotriesterase, (PTE), called SsoPox, able to degrade organophosphate compounds has been previously determined and isolated from Sulfolobus solfataricus and then expressed in Escherichia coli BL21-DE3. In this experimental work a new biotechnological process for the production of SsoPox by using fermentation technologies has been studied. First of all, diverse physiological settings were investigated in shake flasks to optimize growth conditions, enhancing enzyme expression and production by using diverse concentrations and type of inducers. Batch and fed-batch experiments were performed in 2-L scale reactors in order to maximize enzyme production. Besides different extraction methods have been applied to recover the enzyme and thermal treatments coupled to a membrane based downstream (ultrafiltration/diafiltration) have been tuned to achieve a sound purification compatible with diverse applications (e.g. as immobilized enzyme..etc.).