An acetylcholinesterase purified from the greenbug (Schizaphis graminum) with some unique enzymological and pharmacological characteristics.
An acetylcholinesterase (AChE, EC 3.1.1.7) was purified from the greenbug, Schizaphis graminum (Rondani). The maximum velocities (Vmax) for hydrolyzing acetylthiocholine (ATC), acetyl-(beta-methyl) thiocholine (AbetaMTC), propionylthiocholine, and S-butyrylthiocholine were 78.0, 67.0, 37.4, and 2.3 micromol/min/mg, and the Michaelis constants (Km) were 57.6, 60.6, 31.3, and 33.4 microM, respectively. More than 98% of AChE activity was inhibited by 10 microM eserine or BW284C51, but only 7% of the activity was inhibited by ethopropazine at the same concentration. Based on the substrate and inhibitor specificities, the purified enzyme appeared to be a true AChE. Nondenaturing polyacrylamide gel electrophoresis (PAGE) and isoelectric focusing of the purified AChE revealed three molecular forms. The isoelectric points were 7.3 for the major form and 6.3 and 7.1 for two minor forms. The major form of purified AChE showed molecular masses of 129 kDa for its native protein and 72 kDa for its subunits on SDS-PAGE. However, the purified AChE exhibited some distinctive characteristics including: (1) lack of affinity to the affinity ligand 3-(carboxyphenyl) ethyldimethyl ammonium, which has been used widely in purification of AChE from various insect species; and (2) 20-200-fold higher substrate-inhibition thresholds for ATC and AbetaMTC than AChE from other insect species. These biochemical properties may reflect structural differences of AChE purified from the greenbug compared with that from other insect species.
Gao
J R
JR
Department of Entomology, 123 Waters Hall, Kansas State University, Manhattan, KS 66506, USA.
Zhu
K Y
KY
D000110:Acetylcholinesterase / Q000737:chemistry / Q000145:classification / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D001042:Aphids / Q000201:enzymology*; D002800:Cholinesterase Inhibitors / Q000494:pharmacology; D007700:Kinetics; D013379:Substrate Specificity
D002800:Cholinesterase Inhibitors; D000110:Acetylcholinesterase