Quantum dot labeling of butyrylcholinesterase maintains substrate and inhibitor interactions and cell adherence
Nir Waiskopf1, Itzhak Shweky, Itai Lieberman
1The Institute of Chemistry and the Center for Nanoscience and Nanotechnology, The Hebrew University of Jerusalem, Edmond J. Safra Campus, Givat Ram, Jerusalem.
Abstract:
Butyrylcholinesterase (BChE) is the major acetylcholine hydrolyzing enzyme in peripheral mammalian systems. It can either reside in the circulation or adhere to cells and tissues and protect them from anticholinesterases, including insecticides and poisonous nerve gases. In humans, impaired cholinesterase functioning is causally involved in many pathologies, including Alzheimer's and Parkinson's diseases, trait anxiety, and post stroke conditions. Recombinant cholinesterases have been developed for therapeutic use; therefore, it is important to follow their in vivo path, location, and interactions. Traditional labeling methods, such as fluorescent dyes and proteins, generally suffer from sensitivity to environmental conditions, from proximity to different molecules or special enzymes which can alter them, and from relatively fast photobleaching. In contrast, emerging development in synthesis and surface engineering of semiconductor nanocrystals enable their use to detect and follow molecules in biological milieus at high sensitivity and in real time. Therefore, we developed a platform for conjugating highly purified recombinant human BChE dimers (rhBChE) to CdSe/CdZnS quantum dots (QDs). We report the development and characterization of highly fluorescent aqueous soluble QD-rhBChE conjugates, present maintenance of hydrolytic activity, inhibitor sensitivity, and adherence to the membrane of cultured live cells of these conjugates, and outline their advantageous features for diverse biological applications.
