원문정보
초록
영어
The discovery and analysis of interacting proteins are essential for delineating biological networks, the understanding of cellular function and for drug discovery. We have developed a bacterial system for the discovery of interacting proteins that, unlike other 2-hybrid systems, allows for the selection of protein pairs on the basis of affinity or expression. This new technology, called APEx 2-hybrid, relies on the Anchored Periplasmic Expression (APEx)1)
of one protein on the periplasmic side of the inner membrane of Escherichia coli and its interacting partner as a soluble, epitope-tagged, periplasmic protein2). After removal of outer membrane and labelling with fluorescent probe, cells expressing protein pairs exhibiting different affinities can be readily distinguished by flow cytometry. APEx 2-hybrid system coupled with multi-color fluorescence activated cell sorting (FACS) enables protein-protein interaction study in
periplasm of E. coli as well as high thrroughput screening of highly interacting proteins from combinatorial libraries. Various applications of APEx 2-hybrid system toward antibody engineering (affinity/expression maturation of antibody and antibody fragments, disulfide bond-free intrabody etc.) will be presented.