Construction of a linker library with widely controllable flexibility for fusion protein design
0301 basic medicine
Luminescent Proteins
0303 health sciences
03 medical and health sciences
Bacterial Proteins
Protein Conformation
Recombinant Fusion Proteins
Green Fluorescent Proteins
Fluorescence Resonance Energy Transfer
Molecular Dynamics Simulation
Protein Engineering
Artificial Gene Fusion
DOI:
10.1007/s00253-015-6985-3
Publication Date:
2015-09-22T06:11:53Z
AUTHORS (8)
ABSTRACT
Flexibility or rigidity of the linker between two fused proteins is an important parameter that affects the function of fusion proteins. In this study, we constructed a linker library with five elementary units based on the combination of the flexible (GGGGS) and the rigid (EAAAK) units. Molecular dynamics (MD) simulation showed that more rigid units in the linkers lead to more helical conformation and hydrogen bonds, and less distance fluctuation between the N- and C-termini of the linker. The diversity of linker flexibility of the linker library was then studied by fluorescence resonance energy transfer (FRET) of cyan fluorescent protein (CFP)-yellow fluorescent protein (YFP) fusion proteins, which showed that there is a wide range of distribution of the FRET efficiency. Dissipative particle dynamics (DPD) simulation of CFP-YFP with different linkers also gave identical results with that of FRET efficiency analysis, and we further found that the combination manner of the linker peptide had a remarkable effect on the orientation of CFP and YFP domains. Our studies demonstrated that the construction of the linker library with the widely controllable flexibility could provide appropriate linkers with the desirable characteristics to engineer the fusion proteins with the expected functions.
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