Please use this identifier to cite or link to this item: http://scholarbank.nus.edu.sg/handle/10635/13807
Title: Developing sub-proteomic methods for large scale profiling
Authors: TAN LAI PENG
Keywords: Diffential In-Gel Electrophoresis (DIGE), Activity-based probe (ABP), serine hydrolase, caspase, apoptosis
Issue Date: 24-Apr-2004
Source: TAN LAI PENG (2004-04-24). Developing sub-proteomic methods for large scale profiling. ScholarBank@NUS Repository.
Abstract: We have developed a functional proteomic methodology that makes use of a combination of emerging technologies like differential in-gel electrophoresis (DIGE) and activity-based probes (ABP). The probes are made up of fluorophosphonates (FP) which are mechanism-based suicide inhibitors of serine hydrolases. In DIGE, two pools of proteins are labeled with Cy3-FP probe and Cy5-FP probe, respectively. The labeled proteins are quenched, mixed and separated in the same two-dimensional (2D) gel, allowing quantification of differential serine hydrolases expression. This methodology was extended into quantifying differences in the expression of serine hydrolases between normal and apoptotic cells. In another part of the project, we report efficient in vivo labeling of caspases expressed inside apoptotic HeLa cells using fluorescently-labeled or biotinylated, fluoromethylketone (fmk)-containing probe. Preliminary results with these probes indicated that they were highly cell-permeable, and caspase-8 was isolated and identified by biotinylated fmk-containing probe.
URI: http://scholarbank.nus.edu.sg/handle/10635/13807
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