Properties of acetylcholinesterase from Apis mellifera heads

Abstract

AChE (EC 3.1.1.7) from the bee Apis mellifera head was solubilized by different extraction procedures. Buffers containing the non-ionic detergent Lubrol-PX were more efficient for solubilizing the enzyme and exhibited little inactivation effect when compared with Triton X-100-containing buffers. The acetylthiocholine-splitting activity was shown to be that of true AChE on the basis of inhibitor sensitivity and substrate inhibition. The enzyme from Lubrol-containing and detergent-free extracts had an optimum pH for activity of 9 and 8.7, respectively. The optimum temperature for activity was 45°C. The Arrhenius plot showed that the enzyme has an activation energy of 26 ± 0.8 kJ mol −1. Bee head AChE was susceptible to the presence of divalent cations but monovalent cations such as Na + and K + did not induce any inhibitory effect at concentrations up to 1 M. By contrast, they exhibited an activation effect with a maximum at 0.2 M. Histochemical localization for AChE on frozen sections of honeybees confirmed its specificity as true AChE by the use of specific inhibitors such as iso-OMPA, BW284C51 and eserine. The main AChE activity was located in the neuropil appearing as diffused or clumped dots. An AChE activity coating nuclei of neurones and glial cells in the vicinity of the neuropile was detected with Koelle's reaction. Such a distribution suggests different sites for synthesis, storage, transit and secretion for AChE in the brain.