Part:BBa_K2012001

c-di-GMP tandem riboswitche bc3

Three complete c-di-GMP riboswitches (Bc3, Bc4 and Bc5 RNA) with similar structures, which are arranged in tandem to constitute a triple-tandem (Bc3-5 RNA) riboswitch in the 5′-UTR of the cspABCDE mRNA in Bacillus thuringiensis subsp. chinensis CT-43.

Illustration of riboswitch mechanism

As show in the illustration, transcript of riboswitches region would form a hairpin and terminate transcription under low concentration of c-di-GMP.

Schematic of riboswitches

(a)Comparison of Bc3, Bc4, Bc5 terminators. Red rectangle shows the GC rich region of three terminators, respectively. (b) Secondary structure comparisons of Bc3, Bc4 and Bc5 aptamers with Vc2 aptamer. Conserved motifs such as tetra-loop (blue motif in stem P2), tetra-loop receptor (green motif in stem P3) and G·C base pair (C base in stem P2 and G base in stem P3 were drawn in magenta) connecting P2 with P3 were all colored to facilitate comparison. c-di-GMP was drawn in cyan and its interacting bases drawn in red.(Zhou et al., 2016) (c) Multiply local sequence blast of riboswitches’ terminators. Blue rectangle shows U region of riboswitches.

a. Quantification of Bc3 riboswitch. We discovered an interesting phenomenon. With no IPTG induction (background concentration of c-di-GMP), the intensity of 73F exceeds 106F, corresponding to 5.2 folds. While under 0.5mM IPTG induction, the terminator forming efficiency of 73F is 39 folds higher than it is without IPTG. Promoter strength of J23106:J23117=1185:162. This indicates that Bc3 riboswitch is responsible for facilitating downstream expression in E.coli, especially under high c-di-GMP concentration.

Reference:

Zhou, H., Zheng, C., Su, J., Chen, B., Fu, Y., Xie, Y., . . . He, J. (2016). Characterization of a natural triple-tandem c-di-GMP riboswitch and application of the riboswitch-based dual-fluorescence reporter. Sci Rep, 6, 20871. doi:10.1038/srep20871

Sequence and Features

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]