Cloning, mutagenesis, and expression of the acetylcholinesterase gene from a strain of Musca domestica; the change from a drug-resistant to a sensitive enzyme.
Insect acetylcholinesterase (AChE) is known to be a primary target of organophosphorus and carbamate insecticides. However chronic exposure to these chemicals has led to resistance to applied insecticides, due usually to mutation of the AChE gene. Analysis of the AChE gene (hm) of Musca domestica (the housefly), which is cloned in this report, reveals the relationship between mutation and insecticide resistance. The 2,076 bp hm encodes a mature protein of 612 amino acids (67 kDa), and an 80 residue signal peptide. Unlike the enzyme of 'sensitive' strains, the AChE used in this study was resistant to the organophosphorus insecticide, trichlorphon. DNA sequencing showed that this AChE is identical to that of the sensitive strains with the exception of three amino acids Met-82, Ala-262, and Tyr-327. Site-directed mutagenesis of the Ala-262 and Tyr-327 residues largely restored sensitivity to the insecticide, suggesting that these two residues are the key structural elements controlling sensitivity. In addition to these residues, Glu-234 and Ala-236 in the conserved sequence FGESAG are thought to play a role in modulating sensitivity to organophosphorus insecticides.
Kim
Chung-Sei
CS
School of Agricultural Biotechnology, Seoul National University, Suwon 441-744, Korea.
Kim
Won-Tae
WT
Boo
Kyung-Saeng
KS
Kim
Su-Il
SI
D000110:Acetylcholinesterase / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002800:Cholinesterase Inhibitors / Q000494:pharmacology*; D006793:Houseflies / Q000201:enzymology / Q000235:genetics*; D006863:Hydrogen-Ion Concentration; D007305:Insecticide Resistance / Q000235:genetics; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics; D017354:Point Mutation; D016415:Sequence Alignment; D013696:Temperature; D014236:Trichlorfon / Q000494:pharmacology*
D002800:Cholinesterase Inhibitors; D014236:Trichlorfon; D000110:Acetylcholinesterase