Hybrid sol-gel bio-films: influence of synthetic parameters on behaviour and performance of entrapped His6-tagged organophosphorus hydrolase

The presented work describes the optimising of sol-gel process parameters including the aging time of the sol, the water/silane ratios, type of precursor [tetramethoxysilane (TMOS) or tetraethoxysilane (TEOS)], and the precursor ratios using organically modified silanes [methyltrimethoxysilane (MTMOS) and 3-glycidoxypropyltrimethoxysilane (GPTMS)], for the successful entrapment of hexahistidine-tagged organophosphorus hydrolase (His(6)-OPH) and its applicability during organophosphate detoxification processes. The prepared bio-composite films retained enzyme activity by up to 90 % with expected differences between the starting sols' compositions. The obtained values of K-M and V-max for TMOS/MTMOS and TEOS/GPTMS bio-composites equalled 1.5 +/- A 0.1, 1.4 +/- A 0.3 mM, 0.40 +/- A 0.02 and 1.8 +/- A 0.2 mu M/s for paraoxon, respectively, thus assuming effective application of such biocatalysts during the detoxifications of organophosphorus compounds. Furthermore, the bio-composite films showed enhanced pH and temperature stability with respect to the free enzymes as they can be easily separated from the reactive media, as well as having multiple reusages. Bio-composite materials based on hexahistidine-tagged organophosphorus hydrolase were developed by enzyme encapsulation into silica sol-gel matrixes (tetramethoxysilane/methyltrimethoxysilane, tetraethoxysilane/3-glycidoxypropyltrimethoxysilane). The process of biomaterial production was optimised in several pathways and biomaterials were characterised via IR-spectroscopy, SEM-imaging, so as their catalytic efficiency in organophosphate hydrolysis.