|TUCKOW, ALEXANDER - Former ARS Employee|
|BLOOMQUIST, JEFFREY - University Of Florida|
|Perez De Leon, Adalberto - Beto|
Submitted to: Society for Vector Ecology Bulletin
Publication Type: Abstract Only
Publication Acceptance Date: 8/13/2014
Publication Date: N/A
Technical Abstract: Phlebotomine sand flies are small hematophagous vectors of human and zoonotic leishmaniases present throughout tropical and subtropical areas of the world. These flies present serious problems for military operations and resident populations in the Middle East and other areas where they are endemic. We identified and expressed recombinant acetylcholinesterase 1 (rPpAChE1) of Phlebotomus papatasi, a principal vector of human cutaneous leishmaniasis in the Eastern Hemisphere. The enzyme shared substantial sequence identity with AChE of Lutzomyia spp., the New World Sand Fly, as well as AChE1 of important mosquito species. We hypothesized that PpAChE1 with a single amino acid substitution orthologous to that producing high level organophosphate insensitivity in mosquito AChE1 may generate a rPpAChE1 insensitive to inhibition. We utilized targeted mutagenesis to generate and express the recombinant altered enzyme in a baculovirus expression system. Biochemical properties of the rPpAChE1 containing a substitution of serine for glycine at amino acid position 256 (GenBank accession AFP20868) were quite different from the wild type enzyme, including increase in the Michaelis–Menten constant (Km) requiring higher concentration of substrate to achieve maximal velocity, and vastly decreased sensitivity to inhibition by tested inhibitors.