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MCherry [1-10] is a part of bimolecular fluorescence complementation (BiFC) system.

MCherry [1-10] is a part of bimolecular fluorescence complementation (BiFC) system which could visualize protein interactions. The combination of mCherry [1-10] and mCherry [11] can form the fluorescence of mCherry. With its advantages of a short maturation time and brilliant fluorescence, the mCherry BiFC system could find particular applications for analyzing protein–protein interactions especially in living cells ^[1].

Usage and Biology

We devised the fusion expression of mCherry[1-10] with spNW15, SpyCatcher and SdyCatcher (namely SCSdC-mCh[1-10]). We used mCherry[1-10] and mCherry[11] parts to emit fluorescence to verify the success of protien connection. If SCSdC-mCh[1-10] can connect to SnTST- mCh[11] successfully, the mCherry[1-10] and mCherry[11] parts can emit fluorescence as a characterization.

We also employed AlphaFold to predict the structure of the protein constituted by SpyCatcher, mCherry[1-10], spNW15, mCherry[11] and SpyTag, and discovered that following the successful conjugation of SpyTag and SpyCatcher, mCherry[1-10] and mCherry[11] were successfully complemented.(Figure 1).

Characterization

We used SDS-PAGE to test whether the protein containing mCherry[1-10] had been expressed successfully(Figure 2). The molecular weight of SCSdC-mCh[1-10] (containing mCherry[1-10]) is 76.3 kDa. And we purified the target protein with a molecular weight of approximately 70-100 kDa (Lane 4 - Lane 5), which demonstrated that we had successfully expressed the protein containing mCherry[1-10].

Furthermore, we employed a Fluorescent Inverted microscope to examine whether mCherry[1-10] successfully complemented mCherry[11] and emitted fluorescence (Figure 3). We observed the red fluorescence of mCherry under the Fluorescent Inverted microscope, demonstrating that they functioned successfully.

Sequence and Features

1. ↑ Fan, J., et al., Split mCherry as a new red bimolecular fluorescence complementation system for visualizing protein–protein interactions in living cells. Biochemical and Biophysical Research Communications, 2008. 367(1): p. 47-53.