Characterization of molecular and enzymatic properties of three cholinesterases from the common bed bug, Cimex lectularius

Cimex lectularius;molecular form;acetlycholinesterase;Baculovirus expression;Kinetics;Organophosphate;Carbamate;Salivary gland cholinesterase;Immunohistochemistry;630

농업생명과학대학 농생명공학부 ;

University:서울대학교 대학원

关键词: Cimex lectularius;    molecular form;    acetlycholinesterase;    Baculovirus expression;    Kinetics;    Organophosphate;    Carbamate;    Salivary gland cholinesterase;    Immunohistochemistry;    630;   

Others  :  http://s-space.snu.ac.kr/bitstream/10371/125853/1/000000018433.pdf

美国|英语

来源: Seoul National University Open Repository

【 摘 要 】

ClAChE1, ClAChE2 and ClSChE) from the common bed bug, Cimex lectularius. As determined by activity staining and Western blotting after Native polyacrylamide gel electrophoresis, ClAChE1 was the catalytically main enzyme and abundantly expressed in various tissues whereas ClAChE2 existed in central nervous system (CNS). Both ClAChEs existed in dimeric form connected by a disulfide bridge and were attached to the membrane via a glycophosphatidylinisitol-anchor. To investigate enzymatic properties, three ChEs were functionally expressed using baculovirus expression system. Kinetic analysis using in vitro expressed ClAChEs demonstrated that ClAChE1 had higher catalytic efficiency toward acetylcholine, supporting that ClAChE1 plays a major role in postsynaptic transmission. ClAChE2 showed higher catalytic efficiency toward butyrylcholine, wider substrate spectrum and selective inhibition by iso-OMPA. Inhibition assay using in vitro expressed ClAChEs revealed that ClAChE1 was generally more sensitive to insecticides. The relatively higher correlation between in vitro ClAChE1 inhibition and in vivo toxicity suggests that ClAChE1 is the more relevant toxicological target for organophosphate and carbamate insecticides.ClSChE existed in salivary gland had negligible activity to hydrolyze acetylcholine, however ClSChE function is unrevealed. Under native conditions, ClSChE existed as soluble dimeric form connected by a disulphide bridge or soluble monomeric form. As an Immunohistochemistry of the salivary gland and salivary ducts, it existed as soluble form expressed in cell and passes through lumen. During sucking blood, I confirmed that the bed bug secreted ClSChE through salivary canal in proboscis. In the result of acetylcholine/choline assay, ClSChE has a weak catalytic activity to hydrolyze acetylcholine. Based on the studies, ClSChE has non-neuronal function related to bloodsucking as acetylcholinesterase. These findings are useful in expanding our knowledge on insect AChEs and their evolution.