AI Article Synopsis
- Green fluorescent proteins (GFPs) are essential for monitoring membrane protein properties, but a new coral-derived thermostable GFP (TGP) outperforms conventional jellyfish GFPs in stability and sensitivity.
- TGP can accurately report membrane protein stability at temperatures up to 90 °C, whereas traditional GFPs have lower limits (64 °C and 74 °C).
- The study also introduces high-affinity synthetic nanobodies (sybodies) for TGP, enhancing its utility for membrane protein purification and structural studies.
Article Abstract
Green fluorescent proteins (GFPs) are widely used to monitor membrane protein expression, purification, and stability. An ideal reporter should be stable itself and provide high sensitivity and yield. Here, we demonstrate that a coral (Galaxea fascicularis) thermostable GFP (TGP) is by such reasons an improved tag compared to the conventional jellyfish GFPs. TGP faithfully reports membrane protein stability at temperatures near 90 °C (20-min heating). By contrast, the limit for the two popular GFPs is 64 °C and 74 °C. Replacing GFPs with TGP increases yield for all four test membrane proteins in four expression systems. To establish TGP as an affinity tag for membrane protein purification, several high-affinity synthetic nanobodies (sybodies), including a non-competing pair, are generated, and the crystal structure of one complex is solved. Given these advantages, we anticipate that TGP becomes a widely used tool for membrane protein structural studies.
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| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7729955 | PMC |
| http://dx.doi.org/10.1038/s42003-020-01478-z | DOI Listing |
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