Device

Part:BBa_K5045005:Design

Designed by: Jiayang Liu   Group: iGEM24_BAID-China   (2024-10-01)


AεB ATP sensor


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

To have the strongest ATP sensor's effect as possible, we screened all potential protein pairs in the FPbase to find the pair with maximum quantum yields and Förster radius-the two major factors deciding FRET efficiency-and got the Aquamarine-mBaoJin pair. Also we had to maintain activity of two fluorescent proteins and create a structure that FRET can happen when epsilon subunit is bound with ATP and changes its conformation, thus attached the pair to terminal N and C of epsilon seperately.


Source

The protein sequence of two fluorescents were taken from FPbase: Aquamarine (FPbase 963OH) which is an ECFP derivative, and mBaoJin (FPbase 4XHVQ). Sequence for epsilon subunit was chosen from modified Bacillus subtilis FoF1 ATP synthase in Nano-lantern_ATP1 from 462 to 597 (NCBI AFQ60642.1) slightly modified (597L replaced by F). Nucleotide sequence was the three protein sequences connected and reverse translated.

References