For both research and clinical endeavors, there exists a critical need for an effective method for preserving live mammalian cells. The two most common methods for mammalian cell storage are freezing and drying; however, these methods often cause decreased cell viability. We have developed a strategy for preserving cells that uses the disaccharide trehalose, a naturally- occurring cellular protectant. For trehalose to function properly, it must be delivered to the intracellular environment; however, mammalian cell membranes are impermeable to the hydrophilic sugar trehalose. To overcome this obstacle, we have synthesized hydrophobic trehalose analogs in which the hydrophilic hydroxyl groups of trehalose are masked as esters. Once inside the cell, these ester groups should be cleaved by endogenous esterases, thus releasing free trehalose. To evaluate whether our trehalose analogs are substrates for intracellular esterases, we have developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based esterase assay using porcine liver esterase (PLE), a mammalian esterase, as a model enzyme. We are currently using this assay to evaluate the kinetics of PLE-catalyzed hydrolysis of our trehalose analogs. The results from these assays will help us to better design hydrophobic trehalose analogs for use in cellular protection.<br/>

Kinetic study of porcine liver esterase hydrolysis of cell-permeable analogs of trehalose

251st National Meeting (2016)

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