Part:BBa_K5045005
AεB ATP sensor
AεB (Aquamarine-epsilon-mBaojin) ATP sensor protein consists of three major components: the Aquamarine donor fluorescent protein (BBa_K5045002), the epsilon ATP-binding domain (BBa_K5045001)that is responsible for the specific recognition and binding of ATP molecules, and the mBaojin acceptor fluorescent protein (BBa_K5045003) that provides the fluorescence signal. FRET (Förster resonance energy transfer) is the mechanism by which sensor AεB reflects ATP levels through color changes. The Aquamarine donor, initially in its excited electronic state, may transfer energy to the mBaoJin acceptor through nonradiative dipole-dipole coupling. The efficiency of this energy transfer is inversely proportional to the sixth power of the distance between donor and acceptor, making FRET extremely sensitive to small changes in distance. When ATP binds with the epsilon subunit, the subunit conformational changes occur, causing mBaoJin and Aquamarine to approach each other, so the FRET efficiency increases significantly, and color changes from dark blue to green. The strength of fluorescence changes as the concentration of ATP in the environment changes, as the number of ATP bound ATP sensors changes. By detecting changes in the fluorescence signal, we can monitor the ATP levels in cells in real-time. Further, we can measure the data quantitatively. In this project, we constructed an AεB (Aquamarine-epsilon-mBaojin) ATP sensor plasmid. We inserted the target fragment encoding the AεB ATP sensor into digesting sites of NheI and BamHI in the pCDH-CMV vector by molecular cloning technique. It needs to be inserted into plasmids to be produced in prokaryotic cells or inserted into plasmids and then integrated into cell genomes through lentiviral vectors. We suggest people use A and pCDH-CMV(Addgene #72265) for two purposes separately.
Usage and Biology
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
| None |