Author:
Murakami Agnieszka M.,Nagatomo Katsuhiro,Miyoshi Ichro,Itagaki Shirou,Niwa Yasutaka,Murakami Manabu
Abstract
AbstractWe developed a new method to analyze protein–protein interactions using a dual-inducible prokaryotic expression system. To evaluate protein–protein binding, a chimeric fusion toxin gene was constructed using a DNase-treated short DNA fragment (epitope library) and CcdB, which encodes a DNA topoisomerase II toxin. Protein–protein interactions would affect toxin activity, resulting in colony formation. Using this novel system, we found a new binding site in the voltage-dependent calcium channel α1 subunit (CaV1.2) for the voltage-dependent calcium channel β2 subunit. Prokaryotic expression screening of the β2 subunit using an epitope library of CaV1.2 resulted in two overlapping clones of the C-terminal sequence of CaV1.2. In vitro overlay and immunoprecipitation analyses revealed preferential binding of the C-terminal sequences of CaV1.2 and β2.
Funder
Japan Society for the Promotion of Science
Publisher
Springer Science and Business Media LLC
Cited by
1 articles.
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