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제109회 대한화학회 학술발표회, 총회 및 기기전시회 안내 Site-specific labeling of cell surface proteins with fluorescent probes through Npu_DnaE intein based protein trans-splicing

2012년 2월 23일 19시 22분 05초
BIO.P-707 이곳을 클릭하시면 발표코드에 대한 설명을 보실 수 있습니다.
4월 25일 (수요일) 18:00~21:00
저자 및
민경미, 정덕호, 권영은
동국대학교 의생명공학과, Korea
Here we describe a highly specific, robust and rapid new method for labeling of cell surface proteins with fluorescent probes. The method uses the trans-splicing reaction of Nostoc punctiforme (Npu) DnaE split-intein. Split-intein based protein trans-splicing reaction has been used for site-specific labeling of proteins in vitro for various applications. The trans-splicing reaction is self-processing reaction and can be used for introduction of various synthetic probes to target proteins without a large linker proteins. In this report, we have utilized split-intein based protein trans-splicing reaction for site specific labeling of cell surface proteins. We have prepared a model cell surface protein fused to N-terminal fragment of Npu DnaE Intein and C-terminal fragment of Npu DnaE Intein fused to a fluorescent probe. The model protein was expressed on the cell surface and the proteins were labeled with synthetic fluorescent probes through specific interaction between N- and C-fragment of inteins. This labeling reaction was performed within 30 mins using low concentration of fluorescent molecules (4 uM). No external energy was required for the labeling reaction. This improved labeling method will provide a useful tool for studying functions and mobility of proteins in live cells.