Purification of acethylcholinesterase from the mollusc Mytilus galloprovincialis Lam. and investigation of its kinetic properties

Abstract

The role of acetylcholinesterase in terminating acetylcholine-mediated neurotransmission made it the focus of intense research. In this study the haemolymph Acetylcholinesterase (AChE) from the mollusc Mytilus galloprovincialis was purified to homogeneity by (NH4)(2)SO4 fractional precipitation and affinity chromatography on edrophonium-Sepharose 6B. The enzyme was purified approximatedly 13 fold over the crude extract and was obtained in 3 % yield. The specific activity of purified enzyme was 3 U/ mg protein. It had an optimum pH of 7.5 and showed optimal activity at 35 degrees C. Km and Vmax for acetylthiocholine iodide were 1.3 mM and 0.188 mM/mg/ min, respectively. The purified enzyme migrated as a 51 000 dalton band during polyacrylamide gel electrophoresis under denaturing and reducing conditions. Three methoxyflavones were examined in order to evaluate their potential as anti-Alzheimer's Disease (AD) agents. All of the compounds were shown to be potent AChE inhibitors. Therefore, these compounds may have great value in the development of therapeutic and preventive agents for AD.