Difference between revisions of "Part:BBa K3168003"

Latest revision as of 15:24, 21 October 2019

SmallBitNanoLuc

This basic part codes for the SmallBit of NanoLuc, which can be used as a split reporter. In combination with the LargeBit of NanoLuc and the addition of furimazine (the substrate), bright blue light is emitted (Figure 1). A flexible (GGS)₅ linker is located in front of the SmallBit to enable the formation of fusion proteins. A strep-tag is also included at the end for protein purification. NanoLuc SmallBit variants with different affinities for LargeBit are available (Dixon, 2015). This basic part has a relatively low affinity (2.5*10^-6 M) to prevent high background signals when combined with NanoLuc LargeBit. The affinity can be tuned by incorporating certain mutations (Dixon, 2015).

Figure 1. Split-NanoLuc principle.

Usage and Biology

Split reporters have been used a lot to detect protein-protein interactions and for screening purposes (Dixon, 2015). NanoLuc is a small, structurally robust and very bright luciferase. Many methods have been developed using the split variant of NanoLuc, such as the HiBiT technology and NanoLuc ternary technology (Ohmuro-Matsuyama, 2019).

References

Dixon, A. S., Schwinn, M. K., Hall, M. P., Zimmerman, K., Otto, P., Lubben, T. H., ... & Wood, M. G. (2015). NanoLuc complementation reporter optimized for accurate measurement of protein interactions in cells. ACS chemical biology, 11(2), 400-408.

Ohmuro-Matsuyama, Y., & Ueda, H. (2019). Protein-Protein Interaction Assays Using Split-NanoLuc. In Bioluminescence. IntechOpen.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 85
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

@@ Line 2: / Line 2: @@
 <partinfo>BBa_K3168003 short</partinfo>
-This basic part codes for the small bit of NanoLuc, which can be used as a split reporter. In combination with the large bit of NanoLuc and the addition of furimazine (the substrate), bright blue light is emitted (figure 1). A flexible (GGS)<sub>5</sub> linker is located in front of the small bit to enable the formation of fusion proteins. A strep-tag is also included at the end for protein purification. There are different variants of the NanoLuc small bit with different affinities for the large bit (Dixon, 2015). This basic part has a relatively low affinity (2.5*10<sup>-6</sup>M) to prevent high background signals when combined with NanoLuc large bit. The affinity can be tuned by incorporating certain mutations (Dixon, 2015).
+This basic part codes for the SmallBit of NanoLuc, which can be used as a split reporter. In combination with the LargeBit of NanoLuc and the addition of furimazine (the substrate), bright blue light is emitted (Figure 1). A flexible (GGS)<sub>5</sub> linker is located in front of the SmallBit to enable the formation of fusion proteins. A strep-tag is also included at the end for protein purification. NanoLuc SmallBit variants with different affinities for LargeBit are available (Dixon, 2015). This basic part has a relatively low affinity (2.5*10<sup>-6</sup> M) to prevent high background signals when combined with NanoLuc LargeBit. The affinity can be tuned by incorporating certain mutations (Dixon, 2015).
 [[File:T--TU_Eindhoven--Split-NanoLuc.png|500px|]]