Extreme thermal stability of the antiGFP nanobody – GFP complex

BMC Research Notes

Research Note

Hajnalka Jankovics¹  Balázs Kakasi¹  Eszter Gácsi¹  Ferenc Vonderviszt² 

[1] Bio-Nanosystems Laboratory, Research Institute of Biomolecular and Chemical Engineering, Faculty of Engineering, University of Pannonia, Veszprém, Hungary;Bio-Nanosystems Laboratory, Research Institute of Biomolecular and Chemical Engineering, Faculty of Engineering, University of Pannonia, Veszprém, Hungary;Institute of Technical Physics and Materials Science, Centre for Energy Research, Budapest, Hungary;

关键词: Superfolder GFP;    GFP enhancer nanobody;    Structural stability;    CD;    Fluorescence;   

DOI  :  10.1186/s13104-023-06382-3

 received in 2022-07-30, accepted in 2023-06-07,  发布年份 2023

来源: Springer

【 摘 要 】

ObjectiveThe green fluorescent protein (GFP) and its derivatives are widely used in biomedical research. The manipulation of GFP-tagged proteins by GFP-specific binders, e.g. single-domain antibodies (nanobodies), is of increasing significance. It is therefore important to better understand the properties of antiGFP-GFP interaction in order to establish methodological applications. In this work the interaction of superfolder GFP (sfGFP) and its enhancer nanobody (aGFPenh) was characterized further.ResultsPrevious calorimetric experiments demonstrated that the aGFPenh nanobody binds strongly to sfGFP with a nanomolar affinity. Here we show that this interaction results in a substantial structural stabilization of aGFPenh reflected in a significant increase of its melting temperature by almost 30 °C. The thermal stability of the sfGFP-aGFPenh complex is close to 85 °C in the pH range 7.0–8.5. For therapeutic applications thermoresistance is often an essential factor. Our results suggest that methodologies based on GFP-aGFP interaction can be applied under a wide range of physicochemical conditions. The aGFPenh nanobody seems to be suitable for manipulating sfGFP-labeled targets even in extreme thermophilic organisms.

【 授权许可】

CC BY   
© The Author(s) 2023

【 图 表 】

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