Dos Reis_2024_Int.J.Biol.Macromol_287_138578

We purified acetylcholinesterase from adult Euschistus heros stink bugs (AChEeh) a pest that damages economically important crops by affinity chromatography. An AChEeh inhibitor was bound to the resin, which provided selectivity for the enzyme and yielded 6.82 % of pure AChEeh. We found that the enzyme has specific activity of 115.82 +/- 1.67 U.mg(-1), fold purification of 44.27, molecular weight of approximately 180 kDa on SDS-PAGE and 358.11 +/- 13.84 kDa on size exclusion chromatography under native conditions, and optimal activity at 25 degreesC and pH = 8.0. On the basis of circular dichroism, AChEeh has alpha-helical structure with negative minimum at 208 nm and a positive peak at 193 nm. AChEeh Km and Vmax for acetylthiocholine hydrolysis are 24.33 +/-0.33 microM and 50. +/- 3. U.mg(-1), respectively. We used two inhibitors to evaluate AChEeh specificity: galantamine, a standard AChE inhibitor, and carbofuran, a pesticide, and obtained IC(50) of 102.0 +/- 14.8 and 104.8 +/- 10.71 nM and Ki of 7.00 +/- 2.13 and 8.50 +/- 2.88 nM, respectively. The purified AChEeh could be used to screen selective inhibitors in future screening assays.