Acetylcholinesterase from the charru mussel Mytella charruana: kinetic characterization, physicochemical properties and potential as in vitro biomarker in environmental monitoring of mollusk extraction areas.

Acetylcholinesterase (AChE; EC 3.1.1.7) from aquatic organisms have been used to evaluate the exposure of specimens to pesticides and heavy metals at sublethal levels in environmental samples. AChE of Mytella charruana was extracted to characterize its physicochemical and kinetic properties as well as the effect of organophosphate (dichlorvos, diazinon, chlorpyrifos, methyl-parathion and temephos), carbamates (carbaryl, carbofuran and aldicarb), benzoylureas (diflubenzuron and novaluron), pyrethroid (cypermethrin) and juvenile hormone analog - JHA (pyriproxyfen) and the effect of metal ions: Hg2+, Cd2+, Pb2+, As3+, Cu2+ and Zn2+, in order to evaluate the potential of the enzyme as biomarker. The optimum pH of M. charruana AChE was 8.5 and the maximum activity peak occurred at 48 °C, being highly thermostable maintaining 97.8% of its activity after incubation at 60 °C. The Michaelis-Menten constants (km) for the substrates acetylthiocholine and propionylthiocholine were 2.8 ± 1.26 and 4.94 ± 6.9 mmol·L-1, respectively. The Vmax values for the same substrates were 22.6 ± 0.90 and 10.2 ± 4.94 mU·mg-1, respectively. Specific inhibition results suggest an AChE presenting active site with dimensions between those of AChE and butyrylcholinesterase (BChE). The IC20 values related to the effect of the pesticides on the enzyme showed higher inhibitory power of temephos (0.17 μmol·L-1), followed by aldicarb (0.19 μmol·L-1) and diflubenzuron (0.23 μmol·L-1). Metal ions inhibited M. charruana enzyme in the following order: Hg2+ > Pb2+ > Cd2+ > As3+ > Cu2+ > Zn2+. These data suggest that the enzyme showed potential as in vitro biomarker of the exposure to temephos, mercury, zinc and copper.